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  • Corrigendum: DNA polymerase θ accomplishes translesion synthesis opposite 1,N6-ethenodeoxyadenosine with a remarkably high fidelity in human cells
    Genes Dev. (IF 8.990) Pub Date : 2020-01-01
    Jung-Hoon Yoon; Robert E. Johnson; Louise Prakash; Satya Prakash

    Genes & Development 33: 282–287 (2019)

    更新日期:2020-01-02
  • Reviewers, Volume 33 (2019)
    Genes Dev. (IF 8.990) Pub Date : 2019-12-01

    The editors would like to thank the Editorial Board and the following scientists who reviewed papers and provided advice during 2019.

    更新日期:2019-12-03
  • Erratum: Single-nucleus transcriptomic survey of cell diversity and functional maturation in postnatal mammalian hearts
    Genes Dev. (IF 8.990) Pub Date : 2019-11-01
    Peng Hu; Jian Liu; Juanjuan Zhao; Benjamin J. Wilkins; Katherine Lupino; Hao Wu; Liming Pei

    Genes & Development 32: 1344–1357 (2018)

    更新日期:2019-11-01
  • Erratum: Dedifferentiation by adenovirus E1A due to inactivation of Hippo pathway effectors YAP and TAZ
    Genes Dev. (IF 8.990) Pub Date : 2019-11-01
    Nathan R. Zemke; Dawei Gou; Arnold J. Berk

    Genes & Development 33: 828–843 (2019)

    更新日期:2019-11-01
  • The human nucleolus organizer regions.
    Genes Dev. (IF 8.990) Pub Date : null
    Joseph G Gall

    Although the nucleolus was first described in the early 19th century from both animal and plant cells, human nucleoli and particularly the five human nucleolus organizers have not been well characterized. In this issue of Genes & Development, van Sluis and colleagues (pp. 1688-1701) present a detailed molecular analysis of these organizers, which occur on the short arms of five human chromosomes. The near identity of these arms suggests extensive interchromosomal exchange during evolutionary history.

    更新日期:2019-11-01
  • Reviewers, Volume 33 (2019).
    Genes Dev. (IF 8.990) Pub Date : null

    更新日期:2019-11-01
  • Mixed ubiquitin chains regulate DNA repair.
    Genes Dev. (IF 8.990) Pub Date : null
    Gergely Rona,Michele Pagano

    Diverse linkage in polyubiquitin chain structure gives cells an unparalleled complexity to virtually modulate all aspects of cell biology. Substrates can be covalently modified by ubiquitin chains of different topology. Proper DNA damage response takes advantage of this regulatory system and heavily relies on ubiquitin-based signaling. Moreover, increasing evidence suggests that chain specificity dictates DNA repair outcome. In this issue of Genes & Development, Wu and colleagues (pp. 1702-1717) show that Cezanne and Cezanne2, two paralogous deubiquitinating enzymes that are recruited to sites of DNA damage, ensure proper local polyubiquitin chain composition for downstream DNA repair protein assembly. Their study offers a key insight into the mechanism of crosstalk between linkage-specific ubiquitylation at DNA damage sites, while simultaneously raising important questions for future research.

    更新日期:2019-11-01
  • Paralytic, the Drosophila voltage-gated sodium channel, regulates proliferation of neural progenitors.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-23
    Beverly J Piggott,Christian J Peters,Ye He,Xi Huang,Susan Younger,Lily Yeh Jan,Yuh Nung Jan

    Proliferating cells, typically considered "nonexcitable," nevertheless, exhibit regulation by bioelectric signals. Notably, voltage-gated sodium channels (VGSC) that are crucial for neuronal excitability are also found in progenitors and up-regulated in cancer. Here, we identify a role for VGSC in proliferation of Drosophila neuroblast (NB) lineages within the central nervous system. Loss of paralytic (para), the sole gene that encodes Drosophila VGSC, reduces neuroblast progeny cell number. The type II neuroblast lineages, featuring a population of transit-amplifying intermediate neural progenitors (INP) similar to that found in the developing human cortex, are particularly sensitive to para manipulation. Following a series of asymmetric divisions, INPs normally exit the cell cycle through a final symmetric division. Our data suggests that loss of Para induces apoptosis in this population, whereas overexpression leads to an increase in INPs and overall neuroblast progeny cell numbers. These effects are cell autonomous and depend on Para channel activity. Reduction of Para expression not only affects normal NB development, but also strongly suppresses brain tumor mass, implicating a role for Para in cancer progression. To our knowledge, our studies are the first to identify a role for VGSC in neural progenitor proliferation. Elucidating the contribution of VGSC in proliferation will advance our understanding of bioelectric signaling within development and disease states.

    更新日期:2019-11-01
  • Systematic bromodomain protein screens identify homologous recombination and R-loop suppression pathways involved in genome integrity.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-23
    Jae Jin Kim,Seo Yun Lee,Fade Gong,Anna M Battenhouse,Daniel R Boutz,Aarti Bashyal,Samantha T Refvik,Cheng-Ming Chiang,Blerta Xhemalce,Tanya T Paull,Jennifer S Brodbelt,Edward M Marcotte,Kyle M Miller

    Bromodomain proteins (BRD) are key chromatin regulators of genome function and stability as well as therapeutic targets in cancer. Here, we systematically delineate the contribution of human BRD proteins for genome stability and DNA double-strand break (DSB) repair using several cell-based assays and proteomic interaction network analysis. Applying these approaches, we identify 24 of the 42 BRD proteins as promoters of DNA repair and/or genome integrity. We identified a BRD-reader function of PCAF that bound TIP60-mediated histone acetylations at DSBs to recruit a DUB complex to deubiquitylate histone H2BK120, to allowing direct acetylation by PCAF, and repair of DSBs by homologous recombination. We also discovered the bromo-and-extra-terminal (BET) BRD proteins, BRD2 and BRD4, as negative regulators of transcription-associated RNA-DNA hybrids (R-loops) as inhibition of BRD2 or BRD4 increased R-loop formation, which generated DSBs. These breaks were reliant on topoisomerase II, and BRD2 directly bound and activated topoisomerase I, a known restrainer of R-loops. Thus, comprehensive interactome and functional profiling of BRD proteins revealed new homologous recombination and genome stability pathways, providing a framework to understand genome maintenance by BRD proteins and the effects of their pharmacological inhibition.

    更新日期:2019-11-01
  • Activation of invariant natural killer T cells stimulates adipose tissue remodeling via adipocyte death and birth in obesity.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-16
    Jeu Park,Jin Young Huh,Jiyoung Oh,Jong In Kim,Sang Mun Han,Kyung Cheul Shin,Yong Geun Jeon,Sung Sik Choe,Jiyoung Park,Jae Bum Kim

    In obesity, adipose tissue undergoes dynamic remodeling processes such as adipocyte hypertrophy, hypoxia, immune responses, and adipocyte death. However, whether and how invariant natural killer T (iNKT) cells contribute to adipose tissue remodeling are elusive. In this study, we demonstrate that iNKT cells remove unhealthy adipocytes and stimulate the differentiation of healthy adipocytes. In obese adipose tissue, iNKT cells were abundantly found nearby dead adipocytes. FasL-positive adipose iNKT cells exerted cytotoxic effects to eliminate hypertrophic and pro-inflammatory Fas-positive adipocytes. Furthermore, in vivo adipocyte-lineage tracing mice model showed that activation of iNKT cells by alpha-galactosylceramide promoted adipocyte turnover, eventually leading to potentiation of the insulin-dependent glucose uptake ability in adipose tissue. Collectively, our data propose a novel role of adipose iNKT cells in the regulation of adipocyte turnover in obesity.

    更新日期:2019-11-01
  • Dual functions of angiopoietin-like protein 2 signaling in tumor progression and anti-tumor immunity.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-16
    Haruki Horiguchi,Tsuyoshi Kadomatsu,Ryoma Kurahashi,Chiaki Hara,Keishi Miyata,Masaya Baba,Hironobu Osumi,Kazutoyo Terada,Kimi Araki,Toshiyuki Takai,Tomomi Kamba,W Marston Linehan,Toshiro Moroishi,Yuichi Oike

    Angiopoietin-like protein 2 (ANGPTL2) is a secreted glycoprotein homologous to angiopoietins. Previous studies suggest that tumor cell-derived ANGPTL2 has tumor-promoting function. Here, we conducted mechanistic analysis comparing ANGPTL2 function in cancer progression in a murine syngeneic model of melanoma and a mouse model of translocation renal cell carcinoma (tRCC). ANGPTL2 deficiency in tumor cells slowed tRCC progression, supporting a tumor-promoting role. However, systemic ablation of ANGPTL2 accelerated tRCC progression, supporting a tumor-suppressing role. The syngeneic model also demonstrated a tumor-suppressing role of ANGPTL2 in host tumor microenvironmental cells. Furthermore, the syngeneic model showed that PDGFRα+ fibroblasts in the tumor microenvironment express abundant ANGPTL2 and contribute to tumor suppression. Moreover, host ANGPTL2 facilitates CD8+ T-cell cross-priming and enhances anti-tumor immune responses. Importantly, ANGPTL2 activates dendritic cells through PIR-B-NOTCH signaling and enhances tumor vaccine efficacy. Our study provides strong evidence that ANGPTL2 can function in either tumor promotion or suppression, depending on what cell type it is expressed in.

    更新日期:2019-11-01
  • Human NORs, comprising rDNA arrays and functionally conserved distal elements, are located within dynamic chromosomal regions.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-16
    Marjolein van Sluis,Michael Ó Gailín,Joseph G W McCarter,Hazel Mangan,Alice Grob,Brian McStay

    Human nucleolar organizer regions (NORs), containing ribosomal gene (rDNA) arrays, are located on the p-arms of acrocentric chromosomes (HSA13-15, 21, and 22). Absence of these p-arms from genome references has hampered research on nucleolar formation. Previously, we assembled a distal junction (DJ) DNA sequence contig that abuts rDNA arrays on their telomeric side, revealing that it is shared among the acrocentrics and impacts nucleolar organization. To facilitate inclusion into genome references, we describe sequencing the DJ from all acrocentrics, including three versions of HSA21, ∼3 Mb of novel sequence. This was achieved by exploiting monochromosomal somatic cell hybrids containing single human acrocentric chromosomes with NORs that retain functional potential. Analyses revealed remarkable DJ sequence and functional conservation among human acrocentrics. Exploring chimpanzee acrocentrics, we show that "DJ-like" sequences and abutting rDNA arrays are inverted as a unit in comparison to humans. Thus, rDNA arrays and linked DJs represent a conserved functional locus. We provide direct evidence for exchanges between heterologous human acrocentric p-arms, and uncover extensive structural variation between chromosomes and among individuals. These findings lead us to revaluate the molecular definition of NORs, identify novel genomic structural variation, and provide a rationale for the distinctive chromosomal organization of NORs.

    更新日期:2019-11-01
  • The KDM5A/RBP2 histone demethylase represses NOTCH signaling to sustain neuroendocrine differentiation and promote small cell lung cancer tumorigenesis.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-16
    Matthew G Oser,Amin H Sabet,Wenhua Gao,Abhishek A Chakraborty,Anna C Schinzel,Rebecca B Jennings,Raquel Fonseca,Dennis M Bonal,Matthew A Booker,Abdallah Flaifel,Jesse S Novak,Camilla L Christensen,Hua Zhang,Zachary T Herbert,Michael Y Tolstorukov,Elizabeth J Buss,Kwok-Kin Wong,Roderick T Bronson,Quang-De Nguyen,Sabina Signoretti,William G Kaelin

    More than 90% of small cell lung cancers (SCLCs) harbor loss-of-function mutations in the tumor suppressor gene RB1 The canonical function of the RB1 gene product, pRB, is to repress the E2F transcription factor family, but pRB also functions to regulate cellular differentiation in part through its binding to the histone demethylase KDM5A (also known as RBP2 or JARID1A). We show that KDM5A promotes SCLC proliferation and SCLC's neuroendocrine differentiation phenotype in part by sustaining expression of the neuroendocrine transcription factor ASCL1. Mechanistically, we found that KDM5A sustains ASCL1 levels and neuroendocrine differentiation by repressing NOTCH2 and NOTCH target genes. To test the role of KDM5A in SCLC tumorigenesis in vivo, we developed a CRISPR/Cas9-based mouse model of SCLC by delivering an adenovirus (or an adeno-associated virus [AAV]) that expresses Cre recombinase and sgRNAs targeting Rb1, Tp53, and Rbl2 into the lungs of Lox-Stop-Lox Cas9 mice. Coinclusion of a KDM5A sgRNA decreased SCLC tumorigenesis and metastasis, and the SCLCs that formed despite the absence of KDM5A had higher NOTCH activity compared to KDM5A +/+ SCLCs. This work establishes a role for KDM5A in SCLC tumorigenesis and suggests that KDM5 inhibitors should be explored as treatments for SCLC.

    更新日期:2019-11-01
  • Crosstalk between Lys63- and Lys11-polyubiquitin signaling at DNA damage sites is driven by Cezanne.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-09
    Xiao Wu,Shichang Liu,Cari Sagum,Jianji Chen,Rajesh Singh,Apurva Chaturvedi,John R Horton,Tanuja R Kashyap,David Fushman,Xiaodong Cheng,Mark T Bedford,Bin Wang

    The establishment of polyubiquitin conjugates with distinct linkages play important roles in the DNA damage response. Much remains unknown about the regulation of linkage-specific ubiquitin signaling at sites of DNA damage. Here we reveal that Cezanne (also known as Otud7B) deubiquitinating enzyme promotes the recruitment of Rap80/BRCA1-A complex by binding to Lys63-polyubiquitin and targeting Lys11-polyubiquitin. Using a ubiquitin binding domain protein array screen, we identify that the UBA domains of Cezanne and Cezanne2 (also known as Otud7A) selectively bind to Lys63-linked polyubiquitin. Increased Lys11-linkage ubiquitination due to lack of Cezanne DUB activity compromises the recruitment of Rap80/BRCA1-A. Cezanne2 interacts with Cezanne, facilitating Cezanne in the recruitment of Rap80/BRCA1-A, Rad18, and 53BP1, in cellular resistance to ionizing radiation and DNA repair. Our work presents a model that Cezanne serves as a "reader" of the Lys63-linkage polyubiquitin at DNA damage sites and an "eraser" of the Lys11-linkage ubiquitination, indicating a crosstalk between linkage-specific ubiquitination at DNA damage sites.

    更新日期:2019-11-01
  • Context-specific regulation of cell survival by a miRNA-controlled BIM rheostat.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-09
    Verena Labi,Siying Peng,Filippos Klironomos,Mathias Munschauer,Nicolai Kastelic,Tirtha Chakraborty,Katia Schoeler,Emmanuel Derudder,Manuela Martella,Guido Mastrobuoni,Luis R Hernandez-Miranda,Ines Lahmann,Christine Kocks,Carmen Birchmeier,Stefan Kempa,Leticia Quintanilla-Martinez de Fend,Markus Landthaler,Nikolaus Rajewsky,Klaus Rajewsky

    Knockout of the ubiquitously expressed miRNA-17∼92 cluster in mice produces a lethal developmental lung defect, skeletal abnormalities, and blocked B lymphopoiesis. A shared target of miR-17∼92 miRNAs is the pro-apoptotic protein BIM, central to life-death decisions in mammalian cells. To clarify the contribution of miR-17∼92:Bim interactions to the complex miR-17∼92 knockout phenotype, we used a system of conditional mutagenesis of the nine Bim 3' UTR miR-17∼92 seed matches. Blocking miR-17∼92:Bim interactions early in development phenocopied the lethal lung phenotype of miR-17∼92 ablation and generated a skeletal kinky tail. In the hematopoietic system, instead of causing the predicted B cell developmental block, it produced a selective inability of B cells to resist cellular stress; and prevented B and T cell hyperplasia caused by Bim haploinsufficiency. Thus, the interaction of miR-17∼92 with a single target is essential for life, and BIM regulation by miRNAs serves as a rheostat controlling cell survival in specific physiological contexts.

    更新日期:2019-11-01
  • Cell-type-specific dysregulation of RNA alternative splicing in short tandem repeat mouse knockin models of myotonic dystrophy.
    Genes Dev. (IF 8.990) Pub Date : 2019-10-19
    Curtis A Nutter,Jodi L Bubenik,Ruan Oliveira,Franjo Ivankovic,Łukasz J Sznajder,Benjamin M Kidd,Belinda S Pinto,Brittney A Otero,Helmut A Carter,Eric A Vitriol,Eric T Wang,Maurice S Swanson

    Short tandem repeats (STRs) are prone to expansion mutations that cause multiple hereditary neurological and neuromuscular diseases. To study pathomechanisms using mouse models that recapitulate the tissue specificity and developmental timing of an STR expansion gene, we used rolling circle amplification and CRISPR/Cas9-mediated genome editing to generate Dmpk CTG expansion (CTGexp) knockin models of myotonic dystrophy type 1 (DM1). We demonstrate that skeletal muscle myoblasts and brain choroid plexus epithelial cells are particularly susceptible to Dmpk CTGexp mutations and RNA missplicing. Our results implicate dysregulation of muscle regeneration and cerebrospinal fluid homeostasis as early pathogenic events in DM1.

    更新日期:2019-11-01
  • Evaluating phase separation in live cells: diagnosis, caveats, and functional consequences.
    Genes Dev. (IF 8.990) Pub Date : 2019-10-09
    David T McSwiggen,Mustafa Mir,Xavier Darzacq,Robert Tjian

    The idea that liquid-liquid phase separation (LLPS) may be a general mechanism by which molecules in the complex cellular milieu may self-organize has generated much excitement and fervor in the cell biology community. While this concept is not new, its rise to preeminence has resulted in renewed interest in the mechanisms that shape and drive diverse cellular self-assembly processes from gene expression to cell division to stress responses. In vitro biochemical data have been instrumental in deriving some of the fundamental principles and molecular grammar by which biological molecules may phase separate, and the molecular basis of these interactions. Definitive evidence is lacking as to whether the same principles apply in the physiological environment inside living cells. In this Perspective, we analyze the evidence supporting phase separation in vivo across multiple cellular processes. We find that the evidence for in vivo LLPS is often phenomenological and inadequate to discriminate between phase separation and other possible mechanisms. Moreover, the causal relationship and functional consequences of LLPS in vivo are even more elusive. We underscore the importance of performing quantitative measurements on proteins in their endogenous state and physiological abundance, as well as make recommendations for experiments that may yield more conclusive results.

    更新日期:2019-11-01
  • Role of tumor and host autophagy in cancer metabolism.
    Genes Dev. (IF 8.990) Pub Date : 2019-06-05
    Laura Poillet-Perez,Eileen White

    Macroautophagy (referred to here as autophagy) degrades and recycles cytoplasmic constituents to sustain cellular and mammalian metabolism and survival during starvation. Deregulation of autophagy is involved in numerous diseases, such as cancer. Cancers up-regulate autophagy and depend on it for survival, growth, and malignancy in a tumor cell-autonomous fashion. Recently, it has become apparent that autophagy in host tissues as well as the tumor cells themselves contribute to tumor growth. Understanding how autophagy regulates metabolism and tumor growth has revealed new essential tumor nutrients, where they come from, and how they are supplied and used, which can now be targeted for cancer therapy.

    更新日期:2019-11-01
  • Glioblastoma stem cells: lessons from the tumor hierarchy in a lethal cancer.
    Genes Dev. (IF 8.990) Pub Date : 2019-06-05
    Ryan C Gimple,Shruti Bhargava,Deobrat Dixit,Jeremy N Rich

    Glioblastoma ranks among the most lethal of all human cancers. Glioblastomas display striking cellular heterogeneity, with stem-like glioblastoma stem cells (GSCs) at the apex. Although the original identification of GSCs dates back more than a decade, the purification and characterization of GSCs remains challenging. Despite these challenges, the evidence that GSCs play important roles in tumor growth and response to therapy has grown. Like normal stem cells, GSCs are functionally defined and distinguished from their differentiated tumor progeny at core transcriptional, epigenetic, and metabolic regulatory levels, suggesting that no single therapeutic modality will be universally effective against a heterogenous GSC population. Glioblastomas induce a systemic immunosuppression with mixed responses to oncoimmunologic modalities, suggesting the potential for augmentation of response with a deeper consideration of GSCs. Unfortunately, the GSC literature has been complicated by frequent use of inferior cell lines and a lack of proper functional analyses. Collectively, glioblastoma offers a reliable cancer to study cancer stem cells to better model the human disease and inform improved biologic understanding and design of novel therapeutics.

    更新日期:2019-11-01
  • ATDC is required for the initiation of KRAS-induced pancreatic tumorigenesis.
    Genes Dev. (IF 8.990) Pub Date : 2019-05-03
    Lidong Wang,Huibin Yang,Andrea Zamperone,Daniel Diolaiti,Phillip L Palmbos,Ethan V Abel,Vinee Purohit,Igor Dolgalev,Andrew D Rhim,Mats Ljungman,Christina H Hadju,Christopher J Halbrook,Dafna Bar-Sagi,Marina Pasca di Magliano,Howard C Crawford,Diane M Simeone

    Pancreatic adenocarcinoma (PDA) is an aggressive disease driven by oncogenic KRAS and characterized by late diagnosis and therapeutic resistance. Here we show that deletion of the ataxia-telangiectasia group D-complementing (Atdc) gene, whose human homolog is up-regulated in the majority of pancreatic adenocarcinoma, completely prevents PDA development in the context of oncogenic KRAS. ATDC is required for KRAS-driven acinar-ductal metaplasia (ADM) and its progression to pancreatic intraepithelial neoplasia (PanIN). As a result, mice lacking ATDC are protected from developing PDA. Mechanistically, we show ATDC promotes ADM progression to PanIN through activation of β-catenin signaling and subsequent SOX9 up-regulation. These results provide new insight into PDA initiation and reveal ATDC as a potential target for preventing early tumor-initiating events.

    更新日期:2019-11-01
  • The histone chaperone CAF-1 cooperates with the DNA methyltransferases to maintain Cd4 silencing in cytotoxic T cells.
    Genes Dev. (IF 8.990) Pub Date : 2019-04-13
    Charles Ng,Martin Aichinger,Tung Nguyen,Christy Au,Tariq Najar,Lin Wu,Kai R Mesa,Will Liao,Jean-Pierre Quivy,Benjamin Hubert,Genevieve Almouzni,Johannes Zuber,Dan R Littman

    The transcriptional repression of alternative lineage genes is critical for cell fate commitment. Mechanisms by which locus-specific gene silencing is initiated and heritably maintained during cell division are not clearly understood. To study the maintenance of silent gene states, we investigated how the Cd4 gene is stably repressed in CD8+ T cells. Through CRISPR and shRNA screening, we identified the histone chaperone CAF-1 as a critical component for Cd4 repression. We found that the large subunit of CAF-1, Chaf1a, requires the N-terminal KER domain to associate with the histone deacetylases HDAC1/2 and the histone demethylase LSD1, enzymes that also participate in Cd4 silencing. When CAF-1 was lacking, Cd4 derepression was markedly enhanced in the absence of the de novo DNA methyltransferase Dnmt3a but not the maintenance DNA methyltransferase Dnmt1. In contrast to Dnmt1, Dnmt3a deficiency did not significantly alter levels of DNA methylation at the Cd4 locus. Instead, Dnmt3a deficiency sensitized CD8+ T cells to Cd4 derepression mediated by compromised functions of histone-modifying factors, including the enzymes associated with CAF-1. Thus, we propose that the heritable silencing of the Cd4 gene in CD8+ T cells exploits cooperative functions among the DNA methyltransferases, CAF-1, and histone-modifying enzymes.

    更新日期:2019-11-01
  • Twist2 amplification in rhabdomyosarcoma represses myogenesis and promotes oncogenesis by redirecting MyoD DNA binding.
    Genes Dev. (IF 8.990) Pub Date : 2019-04-13
    Stephen Li,Kenian Chen,Yichi Zhang,Spencer D Barnes,Priscilla Jaichander,Yanbin Zheng,Mohammed Hassan,Venkat S Malladi,Stephen X Skapek,Lin Xu,Rhonda Bassel-Duby,Eric N Olson,Ning Liu

    Rhabdomyosarcoma (RMS) is an aggressive pediatric cancer composed of myoblast-like cells. Recently, we discovered a unique muscle progenitor marked by the expression of the Twist2 transcription factor. Genomic analyses of 258 RMS patient tumors uncovered prevalent copy number amplification events and increased expression of TWIST2 in fusion-negative RMS. Knockdown of TWIST2 in RMS cells results in up-regulation of MYOGENIN and a decrease in proliferation, implicating TWIST2 as an oncogene in RMS. Through an inducible Twist2 expression system, we identified Twist2 as a reversible inhibitor of myogenic differentiation with the remarkable ability to promote myotube dedifferentiation in vitro. Integrated analysis of genome-wide ChIP-seq and RNA-seq data revealed the first dynamic chromatin and transcriptional landscape of Twist2 binding during myogenic differentiation. During differentiation, Twist2 competes with MyoD at shared DNA motifs to direct global gene transcription and repression of the myogenic program. Additionally, Twist2 shapes the epigenetic landscape to drive chromatin opening at oncogenic loci and chromatin closing at myogenic loci. These epigenetic changes redirect MyoD binding from myogenic genes toward oncogenic, metabolic, and growth genes. Our study reveals the dynamic interplay between two opposing transcriptional regulators that control the fate of RMS and provides insight into the molecular etiology of this aggressive form of cancer.

    更新日期:2019-11-01
  • Structural basis for inhibition of a response regulator of σS stability by a ClpXP antiadaptor.
    Genes Dev. (IF 8.990) Pub Date : 2019-04-13
    Victoria Dorich,Christiane Brugger,Arti Tripathi,Joel R Hoskins,Song Tong,Margaret M Suhanovsky,Amita Sastry,Sue Wickner,Susan Gottesman,Alexandra M Deaconescu

    The stationary phase promoter specificity subunit σS (RpoS) is delivered to the ClpXP machinery for degradation dependent on the adaptor RssB. This adaptor-specific degradation of σS provides a major point for regulation and transcriptional reprogramming during the general stress response. RssB is an atypical response regulator and the only known ClpXP adaptor that is inhibited by multiple but dissimilar antiadaptors (IraD, IraP, and IraM). These are induced by distinct stress signals and bind to RssB in poorly understood manners to achieve stress-specific inhibition of σS turnover. Here we present the first crystal structure of RssB bound to an antiadaptor, the DNA damage-inducible IraD. The structure reveals that RssB adopts a compact closed architecture with extensive interactions between its N-terminal and C-terminal domains. The structural data, together with mechanistic studies, suggest that RssB plasticity, conferred by an interdomain glutamate-rich flexible linker, is critical for regulation of σS degradation. Structural modulation of interdomain linkers may thus constitute a general strategy for tuning response regulators.

    更新日期:2019-11-01
  • Ccr4-Not maintains genomic integrity by controlling the ubiquitylation and degradation of arrested RNAPII.
    Genes Dev. (IF 8.990) Pub Date : 2019-04-06
    Haoyang Jiang,Marley Wolgast,Laura M Beebe,Joseph C Reese

    The Ccr4-Not complex regulates essentially every aspect of gene expression, from mRNA synthesis to protein destruction. The Not4 subunit of the complex contains an E3 RING domain and targets proteins for ubiquitin-dependent proteolysis. Ccr4-Not associates with elongating RNA polymerase II (RNAPII), which raises the possibility that it controls the degradation of elongation complex components. Here, we demonstrate that Ccr4-Not controls the ubiquitylation and turnover of Rpb1, the largest subunit of RNAPII, during transcription arrest. Deleting NOT4 or mutating its RING domain strongly reduced the DNA damage-dependent ubiquitylation and destruction of Rpb1. Surprisingly, in vitro ubiquitylation assays indicate that Ccr4-Not does not directly ubiquitylate Rpb1 but instead promotes Rpb1 ubiquitylation by the HECT domain-containing ligase Rsp5. Genetic analyses suggest that Ccr4-Not acts upstream of RSP5, where it acts to initiate the destruction process. Ccr4-Not binds Rsp5 and forms a ternary complex with it and the RNAPII elongation complex. Analysis of mutant Ccr4-Not lacking the RING domain of Not4 suggests that it both recruits Rsp5 and delivers the E2 Ubc4/5 to RNAPII. Our work reveals a previously unknown function of Ccr4-Not and identifies an essential new regulator of RNAPII turnover during genotoxic stress.

    更新日期:2019-11-01
  • The long noncoding RNA Falcor regulates Foxa2 expression to maintain lung epithelial homeostasis and promote regeneration.
    Genes Dev. (IF 8.990) Pub Date : 2019-03-30
    Daniel T Swarr,Michael Herriges,Shanru Li,Mike Morley,Sharlene Fernandes,Anusha Sridharan,Su Zhou,Benjamin A Garcia,Kathleen Stewart,Edward E Morrisey

    Transcription factors (TFs) are dosage-sensitive master regulators of gene expression, with haploinsufficiency frequently leading to life-threatening disease. Numerous mechanisms have evolved to tightly regulate the expression and activity of TFs at the transcriptional, translational, and posttranslational levels. A subset of long noncoding RNAs (lncRNAs) is spatially correlated with transcription factors in the genome, but the regulatory relationship between these lncRNAs and their neighboring TFs is unclear. We identified a regulatory feedback loop between the TF Foxa2 and a downstream lncRNA, Falcor (Foxa2-adjacent long noncoding RNA). Foxa2 directly represses Falcor expression by binding to its promoter, while Falcor functions in cis to positively regulate the expression of Foxa2. In the lung, loss of Falcor is sufficient to lead to chronic inflammatory changes and defective repair after airway epithelial injury. Moreover, disruption of the Falcor-Foxa2 regulatory feedback loop leads to altered cell adhesion and migration, in turn resulting in chronic peribronchial airway inflammation and goblet cell metaplasia. These data reveal that the lncRNA Falcor functions within a regulatory feedback loop to fine-tune the expression of Foxa2, maintain airway epithelial homeostasis, and promote regeneration.

    更新日期:2019-11-01
  • Structural basis of recognition and destabilization of the histone H2B ubiquitinated nucleosome by the DOT1L histone H3 Lys79 methyltransferase.
    Genes Dev. (IF 8.990) Pub Date : 2019-03-30
    Seongmin Jang,Chanshin Kang,Han-Sol Yang,Taeyang Jung,Hans Hebert,Ka Young Chung,Seung Joong Kim,Sungchul Hohng,Ji-Joon Song

    DOT1L is a histone H3 Lys79 methyltransferase whose activity is stimulated by histone H2B Lys120 ubiquitination, suggesting cross-talk between histone H3 methylation and H2B ubiquitination. Here, we present cryo-EM structures of DOT1L complexes with unmodified or H2B ubiquitinated nucleosomes, showing that DOT1L recognizes H2B ubiquitin and the H2A/H2B acidic patch through a C-terminal hydrophobic helix and an arginine anchor in DOT1L, respectively. Furthermore, the structures combined with single-molecule FRET experiments show that H2B ubiquitination enhances a noncatalytic function of the DOT1L-destabilizing nucleosome. These results establish the molecular basis of the cross-talk between H2B ubiquitination and H3 Lys79 methylation as well as nucleosome destabilization by DOT1L.

    更新日期:2019-11-01
  • Hippo signaling does it again: arbitrating cardiac fibroblast identity and activation.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-05
    Anne Katrine Z Johansen,Jeffery D Molkentin

    The Hippo pathway is an evolutionarily conserved kinase cascade that is fundamental for tissue development, homeostasis, and regeneration. In the developing mammalian heart, Hippo signaling regulates cardiomyocyte numbers and organ size. While cardiomyocytes in the adult heart are largely postmitotic, Hippo deficiency can increase proliferation of these cells and affect cardiac regenerative capacity. Recent studies have also shown that resident cardiac fibroblasts play a critical role in disease responsiveness and healing, and in this issue of Genes and Development, Xiao and colleagues (pp. 1491-1505) demonstrate that Hippo signaling also integrates the activity of fibroblasts in the heart. They show that Hippo signaling normally maintains the cardiac fibroblast in a resting state and, conversely, its inactivation during disease-related stress results in a spontaneous transition toward a myofibroblast state that underlies fibrosis and ventricular remodeling. This phenotypic switch is associated with increased cytokine signaling that promotes nonautonomous resident fibroblast and myeloid cell activation.

    更新日期:2019-11-01
  • NBA1, a new player in the Brca1 A complex, is required for DNA damage resistance and checkpoint control.
    Genes Dev. (IF 8.990) Pub Date : 2009-03-06
    Bin Wang,Kristen Hurov,Kay Hofmann,Stephen J Elledge

    The ability to sense and respond to DNA damage is critical to maintenance of genomic stability and the prevention of cancer. In this study, we employed a genetic screen to identify a gene, NBA1 (new component of the BRCA1 A complex), that is required for resistance to ionizing radiation. The NBA1 protein localizes to sites of DNA damage and is required for G2/M checkpoint control. Proteomic analysis revealed that NBA1 is a component of the BRCA1 A complex, which also contains Brca1/Bard1, Abra1, RAP80, BRCC36, and BRE. NBA1 is required to maintain BRE and Abra1 abundance and for the recruitment of BRCA1 to sites of DNA damage. In depth bioinformatics analysis revealed that the BRCA1 A complex bears striking similarities to the 19S proteasome complex. Furthermore, we show that four members of the BRCA1-A complex possess a polyubiquitin chain-binding capability, thus forming a complex that might facilitate the deubiquitinating activity of the deubiquitination enzyme BRCC36 or the E3 ligase activity of the BRCA1/BARD1 ligase. These findings provide a new perspective from which to view the BRCA1 A complex.

    更新日期:2019-11-01
  • MERIT40 facilitates BRCA1 localization and DNA damage repair.
    Genes Dev. (IF 8.990) Pub Date : 2009-03-06
    Lin Feng,Jun Huang,Junjie Chen

    The product of breast cancer susceptibility gene 1, BRCA1, plays pivotal roles in the maintenance of genomic integrity. Mounting evidence indicates that BRCA1 associates with many proteins or protein complexes to regulate diverse processes important for the cellular response to DNA damage. One of these complexes, which mediates the accumulation of BRCA1 at sites of DNA breaks, involves the ubiquitin-binding motif (UIM)-containing protein RAP80, a coiled-coil domain protein CCDC98/Abraxas, and a deubiquitinating enzyme BRCC36. Here we describe the characterization of a novel component of this complex, MERIT40 (Mediator of Rap80 Interactions and Targeting 40 kd), which together with an adaptor protein BRE/BRCC45, enforces the BRCA1-dependent DNA damage response. MERIT40 is assembled into this RAP80/CCDC98-containing complex via its direct interaction with BRE/BRCC45. Importantly, MERIT40 regulates BRCA1 retention at DNA breaks and checkpoint function primarily via a role in maintaining the stability of BRE and this five-subunit protein complex at sites of DNA damage. Together, our study reveals that a stable complex containing MERIT40 acts early in DNA damage response and regulates damage-dependent BRCA1 localization.

    更新日期:2019-11-01
  • A novel, mitogen-activated nuclear kinase is related to a Drosophila developmental regulator.
    Genes Dev. (IF 8.990) Pub Date : 1996-02-01
    G V Denis,M R Green

    Although the ultimate targets of many signal transduction pathways are nuclear transcription factors, the vast majority of known protein kinases are cytosolic. Here, we report on a novel human kinase that is present exclusively in the nucleus. Kinase activity is increased upon cellular proliferation and is markedly elevated in patients with acute and chronic lymphocytic leukemias. We have identified a human gene that encodes this nuclear kinase and find that it is closely related to Drosophila female sterile homeotic (fsh), a developmental regulator with no known biochemical activity. Collectively, these results suggest that this nuclear kinase is a component of a signal transduction pathway that plays a role in Drosophila development and human growth control.

    更新日期:2019-11-01
  • Human stem cell models: lessons for pancreatic development and disease.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-05
    Bjoern Gaertner,Andrea C Carrano,Maike Sander

    A comprehensive understanding of mechanisms that underlie the development and function of human cells requires human cell models. For the pancreatic lineage, protocols have been developed to differentiate human pluripotent stem cells (hPSCs) into pancreatic endocrine and exocrine cells through intermediates resembling in vivo development. In recent years, this differentiation system has been employed to decipher mechanisms of pancreatic development, congenital defects of the pancreas, as well as genetic forms of diabetes and exocrine diseases. In this review, we summarize recent insights gained from studies of pancreatic hPSC models. We discuss how genome-scale analyses of the differentiation system have helped elucidate roles of chromatin state, transcription factors, and noncoding RNAs in pancreatic development and how the analysis of cells with disease-relevant mutations has provided insight into the molecular underpinnings of genetically determined diseases of the pancreas.

    更新日期:2019-11-01
  • Metabolic dependencies and vulnerabilities in leukemia.
    Genes Dev. (IF 8.990) Pub Date : 2019-11-05
    Marissa Rashkovan,Adolfo Ferrando

    Leukemia cell proliferation requires up-regulation and rewiring of metabolic pathways to feed anabolic cell growth. Oncogenic drivers directly and indirectly regulate metabolic pathways, and aberrant metabolism is central not only for leukemia proliferation and survival, but also mediates oncogene addiction with significant implications for the development of targeted therapies. This review explores leukemia metabolic circuitries feeding anabolism, redox potential, and energy required for tumor propagation with an emphasis on emerging therapeutic opportunities.

    更新日期:2019-11-01
  • Checkpoint inhibition of origin firing prevents DNA topological stress.
    Genes Dev. (IF 8.990) Pub Date : 2019-10-19
    Esther C Morafraile,Christine Hänni,George Allen,Theresa Zeisner,Caroline Clarke,Mark C Johnson,Miguel M Santos,Lauren Carroll,Nicola E Minchell,Jonathan Baxter,Peter Banks,Dave Lydall,Philip Zegerman

    A universal feature of DNA damage and replication stress in eukaryotes is the activation of a checkpoint-kinase response. In S-phase, the checkpoint inhibits replication initiation, yet the function of this global block to origin firing remains unknown. To establish the physiological roles of this arm of the checkpoint, we analyzed separation of function mutants in the budding yeast Saccharomyces cerevisiae that allow global origin firing upon replication stress, despite an otherwise normal checkpoint response. Using genetic screens, we show that lack of the checkpoint-block to origin firing results in a dependence on pathways required for the resolution of topological problems. Failure to inhibit replication initiation indeed causes increased DNA catenation, resulting in DNA damage and chromosome loss. We further show that such topological stress is not only a consequence of a failed checkpoint response but also occurs in an unperturbed S-phase when too many origins fire simultaneously. Together we reveal that the role of limiting the number of replication initiation events is to prevent DNA topological problems, which may be relevant for the treatment of cancer with both topoisomerase and checkpoint inhibitors.

    更新日期:2019-11-01
  • Hi-C guided assemblies reveal conserved regulatory topologies on X and autosomes despite extensive genome shuffling.
    Genes Dev. (IF 8.990) Pub Date : 2019-10-12
    Gina Renschler,Gautier Richard,Claudia Isabelle Keller Valsecchi,Sarah Toscano,Laura Arrigoni,Fidel Ramírez,Asifa Akhtar

    Genome rearrangements that occur during evolution impose major challenges on regulatory mechanisms that rely on three-dimensional genome architecture. Here, we developed a scaffolding algorithm and generated chromosome-length assemblies from Hi-C data for studying genome topology in three distantly related Drosophila species. We observe extensive genome shuffling between these species with one synteny breakpoint after approximately every six genes. A/B compartments, a set of large gene-dense topologically associating domains (TADs), and spatial contacts between high-affinity sites (HAS) located on the X chromosome are maintained over 40 million years, indicating architectural conservation at various hierarchies. Evolutionary conserved genes cluster in the vicinity of HAS, while HAS locations appear evolutionarily flexible, thus uncoupling functional requirement of dosage compensation from individual positions on the linear X chromosome. Therefore, 3D architecture is preserved even in scenarios of thousands of rearrangements highlighting its relevance for essential processes such as dosage compensation of the X chromosome.

    更新日期:2019-11-01
  • Structural basis for distinct roles of SMAD2 and SMAD3 in FOXH1 pioneer-directed TGF-β signaling.
    Genes Dev. (IF 8.990) Pub Date : 2019-10-05
    Eric Aragón,Qiong Wang,Yilong Zou,Sophie M Morgani,Lidia Ruiz,Zuzanna Kaczmarska,Jie Su,Carles Torner,Lin Tian,Jing Hu,Weiping Shu,Saloni Agrawal,Tiago Gomes,José A Márquez,Anna-Katerina Hadjantonakis,Maria J Macias,Joan Massagué

    TGF-β receptors phosphorylate SMAD2 and SMAD3 transcription factors, which then form heterotrimeric complexes with SMAD4 and cooperate with context-specific transcription factors to activate target genes. Here we provide biochemical and structural evidence showing that binding of SMAD2 to DNA depends on the conformation of the E3 insert, a structural element unique to SMAD2 and previously thought to render SMAD2 unable to bind DNA. Based on this finding, we further delineate TGF-β signal transduction by defining distinct roles for SMAD2 and SMAD3 with the forkhead pioneer factor FOXH1 as a partner in the regulation of differentiation genes in mouse mesendoderm precursors. FOXH1 is prebound to target sites in these loci and recruits SMAD3 independently of TGF-β signals, whereas SMAD2 remains predominantly cytoplasmic in the basal state and set to bind SMAD4 and join SMAD3:FOXH1 at target promoters in response to Nodal TGF-β signals. The results support a model in which signal-independent binding of SMAD3 and FOXH1 prime mesendoderm differentiation gene promoters for activation, and signal-driven SMAD2:SMAD4 binds to promoters that are preloaded with SMAD3:FOXH1 to activate transcription.

    更新日期:2019-11-01
  • Termination of pre-mRNA splicing requires that the ATPase and RNA unwindase Prp43p acts on the catalytic snRNA U6.
    Genes Dev. (IF 8.990) Pub Date : 2019-09-29
    Rebecca Toroney,Klaus H Nielsen,Jonathan P Staley

    The termination of pre-mRNA splicing functions to discard suboptimal substrates, thereby enhancing fidelity, and to release excised introns in a manner coupled to spliceosome disassembly, thereby allowing recycling. The mechanism of termination, including the RNA target of the DEAH-box ATPase Prp43p, remains ambiguous. We discovered a critical role for nucleotides at the 3' end of the catalytic U6 small nuclear RNA in splicing termination. Although conserved sequence at the 3' end is not required, 2' hydroxyls are, paralleling requirements for Prp43p biochemical activities. Although the 3' end of U6 is not required for recruiting Prp43p to the spliceosome, the 3' end cross-links directly to Prp43p in an RNA-dependent manner. Our data indicate a mechanism of splicing termination in which Prp43p translocates along U6 from the 3' end to disassemble the spliceosome and thereby release suboptimal substrates or excised introns. This mechanism reveals that the spliceosome becomes primed for termination at the same stage it becomes activated for catalysis, implying a requirement for stringent control of spliceosome activity within the cell.

    更新日期:2019-11-01
  • Hippo pathway deletion in adult resting cardiac fibroblasts initiates a cell state transition with spontaneous and self-sustaining fibrosis.
    Genes Dev. (IF 8.990) Pub Date : 2019-09-29
    Yang Xiao,Matthew C Hill,Lele Li,Vaibhav Deshmukh,Thomas J Martin,Jun Wang,James F Martin

    Cardiac fibroblasts (CFs) respond to injury by transitioning through multiple cell states, including resting CFs, activated CFs, and myofibroblasts. We report here that Hippo signaling cell-autonomously regulates CF fate transitions and proliferation, and non-cell-autonomously regulates both myeloid and CF activation in the heart. Conditional deletion of Hippo pathway kinases, Lats1 and Lats2, in uninjured CFs initiated a self-perpetuating fibrotic response in the adult heart that was exacerbated by myocardial infarction (MI). Single cell transcriptomics showed that uninjured Lats1/2 mutant CFs spontaneously transitioned to a myofibroblast cell state. Through gene regulatory network reconstruction, we found that Hippo-deficient myofibroblasts deployed a network of transcriptional regulators of endoplasmic reticulum (ER) stress, and the unfolded protein response (UPR) consistent with elevated secretory activity. We observed an expansion of myeloid cell heterogeneity in uninjured Lats1/2 CKO hearts with similarity to cells recovered from control hearts post-MI. Integrated genome-wide analysis of Yap chromatin occupancy revealed that Yap directly activates myofibroblast cell identity genes, the proto-oncogene Myc, and an array of genes encoding pro-inflammatory factors through enhancer-promoter looping. Our data indicate that Lats1/2 maintain the resting CF cell state through restricting the Yap-induced injury response.

    更新日期:2019-11-01
  • Planarians recruit piRNAs for mRNA turnover in adult stem cells.
    Genes Dev. (IF 8.990) Pub Date : 2019-09-21
    Iana V Kim,Elizabeth M Duncan,Eric J Ross,Vladyslava Gorbovytska,Stephanie H Nowotarski,Sarah A Elliott,Alejandro Sánchez Alvarado,Claus-D Kuhn

    PIWI proteins utilize small RNAs called piRNAs to silence transposable elements, thereby protecting germline integrity. In planarian flatworms, PIWI proteins are essential for regeneration, which requires adult stem cells termed neoblasts. Here, we characterize planarian piRNAs and examine the roles of PIWI proteins in neoblast biology. We find that the planarian PIWI proteins SMEDWI-2 and SMEDWI-3 cooperate to degrade active transposons via the ping-pong cycle. Unexpectedly, we discover that SMEDWI-3 plays an additional role in planarian mRNA surveillance. While SMEDWI-3 degrades numerous neoblast mRNAs in a homotypic ping-pong cycle, it is also guided to another subset of neoblast mRNAs by antisense piRNAs and binds these without degrading them. Mechanistically, the distinct activities of SMEDWI-3 are primarily dictated by the degree of complementarity between target mRNAs and antisense piRNAs. Thus, PIWI proteins enable planarians to repurpose piRNAs for potentially critical roles in neoblast mRNA turnover.

    更新日期:2019-11-01
  • The Integrator complex cleaves nascent mRNAs to attenuate transcription.
    Genes Dev. (IF 8.990) Pub Date : 2019-09-19
    Deirdre C Tatomer,Nathan D Elrod,Dongming Liang,Mei-Sheng Xiao,Jeffrey Z Jiang,Michael Jonathan,Kai-Lieh Huang,Eric J Wagner,Sara Cherry,Jeremy E Wilusz

    Cellular homeostasis requires transcriptional outputs to be coordinated, and many events post-transcription initiation can dictate the levels and functions of mature transcripts. To systematically identify regulators of inducible gene expression, we performed high-throughput RNAi screening of the Drosophila Metallothionein A (MtnA) promoter. This revealed that the Integrator complex, which has a well-established role in 3' end processing of small nuclear RNAs (snRNAs), attenuates MtnA transcription during copper stress. Integrator complex subunit 11 (IntS11) endonucleolytically cleaves MtnA transcripts, resulting in premature transcription termination and degradation of the nascent RNAs by the RNA exosome, a complex also identified in the screen. Using RNA-seq, we then identified >400 additional Drosophila protein-coding genes whose expression increases upon Integrator depletion. We focused on a subset of these genes and confirmed that Integrator is bound to their 5' ends and negatively regulates their transcription via IntS11 endonuclease activity. Many noncatalytic Integrator subunits, which are largely dispensable for snRNA processing, also have regulatory roles at these protein-coding genes, possibly by controlling Integrator recruitment or RNA polymerase II dynamics. Altogether, our results suggest that attenuation via Integrator cleavage limits production of many full-length mRNAs, allowing precise control of transcription outputs.

    更新日期:2019-11-01
  • TGIF transcription factors repress acetyl CoA metabolic gene expression and promote intestinal tumor growth.
    Genes Dev. (IF 8.990) Pub Date : 2019-02-28
    Anant Shah,Tiffany A Melhuish,Todd E Fox,Henry F Frierson,David Wotton

    Tgif1 (thymine-guanine-interacting factor 1) and Tgif2 repress gene expression by binding directly to DNA or interacting with transforming growth factor (TGF) β-responsive SMADs. Tgifs are essential for embryogenesis and may function in tumor progression. By analyzing both gain and loss of Tgif function in a well-established mouse model of intestinal cancer, we show that Tgifs promote adenoma growth in the context of mutant Apc (adenomatous polyposis coli). Despite the tumor-suppressive role of TGFβ signaling, transcriptome profiling of colon tumors suggests minimal effect of Tgifs on the TGFβ pathway. Instead, it appears that Tgifs, which are up-regulated in Apc mutant colon tumors, contribute to reprogramming metabolic gene expression. Integrating gene expression data from colon tumors with other gene expression and chromatin-binding data identifies a set of direct Tgif target genes encoding proteins involved in acetyl CoA and pyruvate metabolism. Analysis of both tumor and nontumor tissues indicates that these genes are targets of Tgif repression in multiple settings, suggesting that this is a core Tgif function. We propose that Tgifs play an important role in regulating basic energy metabolism in normal cells, and that this function of Tgifs is amplified in some cancers.

    更新日期:2019-11-01
  • ATR acts stage specifically to regulate multiple aspects of mammalian meiotic silencing.
    Genes Dev. (IF 8.990) Pub Date : 2013-07-05
    Hélène Royo,Haydn Prosser,Yaroslava Ruzankina,Shantha K Mahadevaiah,Jeffrey M Cloutier,Marek Baumann,Tomoyuki Fukuda,Christer Höög,Attila Tóth,Dirk G de Rooij,Allan Bradley,Eric J Brown,James M A Turner

    In mammals, homologs that fail to synapse during meiosis are transcriptionally inactivated. This process, meiotic silencing, drives inactivation of the heterologous XY bivalent in male germ cells (meiotic sex chromosome inactivation [MSCI]) and is thought to act as a meiotic surveillance mechanism. The checkpoint protein ATM and Rad3-related (ATR) localizes to unsynapsed chromosomes, but its role in the initiation and maintenance of meiotic silencing is unknown. Here we show that ATR has multiple roles in silencing. ATR first regulates HORMA (Hop1, Rev7, and Mad2) domain protein HORMAD1/2 phosphorylation and localization of breast cancer I (BRCA1) and ATR cofactors ATR-interacting peptide (ATRIP)/topoisomerase 2-binding protein 1 (TOPBP1) at unsynapsed axes. Later, it acts as an adaptor, transducing signaling at unsynapsed axes into surrounding chromatin in a manner that requires interdependence with mediator of DNA damage checkpoint 1 (MDC1) and H2AFX. Finally, ATR catalyzes histone H2AFX phosphorylation, the epigenetic event leading to gene inactivation. Using a novel genetic strategy in which MSCI is used to silence a chosen gene in pachytene, we show that ATR depletion does not disrupt the maintenance of silencing and that silencing comprises two phases: The first is dynamic and reversible, and the second is stable and irreversible. Our work identifies a role for ATR in the epigenetic regulation of gene expression and presents a new technique for ablating gene function in the germline.

    更新日期:2019-11-01
  • The transcriptional repressor CDP (Cutl1) is essential for epithelial cell differentiation of the lung and the hair follicle.
    Genes Dev. (IF 8.990) Pub Date : 2001-09-07
    T Ellis,L Gambardella,M Horcher,S Tschanz,J Capol,P Bertram,W Jochum,Y Barrandon,M Busslinger

    The mammalian Cutl1 gene codes for the CCAAT displacement protein (CDP), which has been implicated as a transcriptional repressor in diverse processes such as terminal differentiation, cell cycle progression, and the control of nuclear matrix attachment regions. To investigate the in vivo function of Cutl1, we have replaced the C-terminal Cut repeat 3 and homeodomain exons with an in-frame lacZ gene by targeted mutagenesis in the mouse. The CDP-lacZ fusion protein is retained in the cytoplasm and fails to repress gene transcription, indicating that the Cutl1(lacZ) allele corresponds to a null mutation. Cutl1 mutant mice on inbred genetic backgrounds are born at Mendelian frequency, but die shortly after birth because of retarded differentiation of the lung epithelia, which indicates an essential role of CDP in lung maturation. A less pronounced delay in lung development allows Cutl1 mutant mice on an outbred background to survive beyond birth. These mice are growth-retarded and develop an abnormal pelage because of disrupted hair follicle morphogenesis. The inner root sheath (IRS) is reduced, and the transcription of Sonic hedgehog and IRS-specific genes is deregulated in Cutl1 mutant hair follicles, consistent with the specific expression of Cutl1 in the progenitors and cell lineages of the IRS. These data implicate CDP in cell-lineage specification during hair follicle morphogenesis, which resembles the role of the related Cut protein in specifying cell fates during Drosophila development.

    更新日期:2019-11-01
  • Functional inactivation of the IGF-I and insulin receptors in skeletal muscle causes type 2 diabetes.
    Genes Dev. (IF 8.990) Pub Date : 2001-08-04
    A M Fernández,J K Kim,S Yakar,J Dupont,C Hernandez-Sanchez,A L Castle,J Filmore,G I Shulman,D Le Roith

    Peripheral insulin resistance and impaired insulin action are the primary characteristics of type 2 diabetes. The first observable defect in this major disorder occurs in muscle, where glucose disposal in response to insulin is impaired. We have developed a transgenic mouse with a dominant-negative insulin-like growth factor-I receptor (KR-IGF-IR) specifically targeted to the skeletal muscle. Expression of KR-IGF-IR resulted in the formation of hybrid receptors between the mutant and the endogenous IGF-I and insulin receptors, thereby abrogating the normal function of these receptors and leading to insulin resistance. Pancreatic beta-cell dysfunction developed at a relative early age, resulting in diabetes. These mice provide an excellent model to study the molecular mechanisms underlying the development of human type 2 diabetes.

    更新日期:2019-11-01
  • Intricate SUMO-based control of the homologous recombination machinery.
    Genes Dev. (IF 8.990) Pub Date : 2019-10-03
    Nalini Dhingra,Xiaolan Zhao

    The homologous recombination (HR) machinery plays multiple roles in genome maintenance. Best studied in the context of DNA double-stranded break (DSB) repair, recombination enzymes can cleave, pair, and unwind DNA molecules, and collaborate with regulatory proteins to execute multiple DNA processing steps before generating specific repair products. HR proteins also help to cope with problems arising from DNA replication, modulating impaired replication forks or filling DNA gaps. Given these important roles, it is not surprising that each HR step is subject to complex regulation to adjust repair efficiency and outcomes as well as to limit toxic intermediates. Recent studies have revealed intricate regulation of all steps of HR by the protein modifier SUMO, which has been increasingly recognized for its broad influence in nuclear functions. This review aims to connect established roles of SUMO with its newly identified effects on recombinational repair and stimulate further thought on many unanswered questions.

    更新日期:2019-11-01
  • Kidney organoids: accurate models or fortunate accidents.
    Genes Dev. (IF 8.990) Pub Date : 2019-10-03
    Melissa H Little,Alexander N Combes

    There are now many reports of human kidney organoids generated via the directed differentiation of human pluripotent stem cells (PSCs) based on an existing understanding of mammalian kidney organogenesis. Such kidney organoids potentially represent tractable tools for the study of normal human development and disease with improvements in scale, structure, and functional maturation potentially providing future options for renal regeneration. The utility of such organotypic models, however, will ultimately be determined by their developmental accuracy. While initially inferred from mouse models, recent transcriptional analyses of human fetal kidney have provided greater insight into nephrogenesis. In this review, we discuss how well human kidney organoids model the human fetal kidney and how the remaining differences challenge their utility.

    更新日期:2019-11-01
  • Melanoma plasticity and phenotypic diversity: therapeutic barriers and opportunities.
    Genes Dev. (IF 8.990) Pub Date : 2019-10-03
    Florian Rambow,Jean-Christophe Marine,Colin R Goding

    An incomplete view of the mechanisms that drive metastasis, the primary cause of cancer-related death, has been a major barrier to development of effective therapeutics and prognostic diagnostics. Increasing evidence indicates that the interplay between microenvironment, genetic lesions, and cellular plasticity drives the metastatic cascade and resistance to therapies. Here, using melanoma as a model, we outline the diversity and trajectories of cell states during metastatic dissemination and therapy exposure, and highlight how understanding the magnitude and dynamics of nongenetic reprogramming in space and time at single-cell resolution can be exploited to develop therapeutic strategies that capitalize on nongenetic tumor evolution.

    更新日期:2019-11-01
  • USP21 deubiquitinase promotes pancreas cancer cell stemness via Wnt pathway activation.
    Genes Dev. (IF 8.990) Pub Date : 2019-09-07
    Pingping Hou,Xingdi Ma,Qiang Zhang,Chang-Jiun Wu,Wenting Liao,Jun Li,Huamin Wang,Jun Zhao,Xin Zhou,Carolyn Guan,Jeffery Ackroyd,Shan Jiang,Jianhua Zhang,Denise J Spring,Y Alan Wang,Ronald A DePinho

    The ubiquitin-specific protease (USP) family is the largest group of cysteine proteases. Cancer genomic analysis identified frequent amplification of USP21 (22%) in human pancreatic ductal adenocarcinoma (PDAC). USP21 overexpression correlates with human PDAC progression, and enforced expression of USP21 accelerates murine PDAC tumor growth and drives PanIN to PDAC progression in immortalized human pancreatic ductal cells. Conversely, depletion of USP21 impairs PDAC tumor growth. Mechanistically, USP21 deubiquitinates and stabilizes the TCF/LEF transcription factor TCF7, which promotes cancer cell stemness. Our work identifies and validates USP21 as a PDAC oncogene, providing a potential druggable target for this intractable disease.

    更新日期:2019-11-01
  • ZCCHC8, the nuclear exosome targeting component, is mutated in familial pulmonary fibrosis and is required for telomerase RNA maturation.
    Genes Dev. (IF 8.990) Pub Date : 2019-09-07
    Dustin L Gable,Valeriya Gaysinskaya,Christine C Atik,C Conover Talbot,Byunghak Kang,Susan E Stanley,Elizabeth W Pugh,Nuria Amat-Codina,Kara M Schenk,Murat O Arcasoy,Cory Brayton,Liliana Florea,Mary Armanios

    Short telomere syndromes manifest as familial idiopathic pulmonary fibrosis; they are the most common premature aging disorders. We used genome-wide linkage to identify heterozygous loss of function of ZCCHC8, a zinc-knuckle containing protein, as a cause of autosomal dominant pulmonary fibrosis. ZCCHC8 associated with TR and was required for telomerase function. In ZCCHC8 knockout cells and in mutation carriers, genomically extended telomerase RNA (TR) accumulated at the expense of mature TR, consistent with a role for ZCCHC8 in mediating TR 3' end targeting to the nuclear RNA exosome. We generated Zcchc8-null mice and found that heterozygotes, similar to human mutation carriers, had TR insufficiency but an otherwise preserved transcriptome. In contrast, Zcchc8-/- mice developed progressive and fatal neurodevelopmental pathology with features of a ciliopathy. The Zcchc8-/- brain transcriptome was highly dysregulated, showing accumulation and 3' end misprocessing of other low-abundance RNAs, including those encoding cilia components as well as the intronless replication-dependent histones. Our data identify a novel cause of human short telomere syndromes-familial pulmonary fibrosis and uncover nuclear exosome targeting as an essential 3' end maturation mechanism that vertebrate TR shares with replication-dependent histones.

    更新日期:2019-11-01
  • Automethylation of PRC2 promotes H3K27 methylation and is impaired in H3K27M pediatric glioma.
    Genes Dev. (IF 8.990) Pub Date : 2019-09-07
    Chul-Hwan Lee,Jia-Ray Yu,Jeffrey Granat,Ricardo Saldaña-Meyer,Joshua Andrade,Gary LeRoy,Ying Jin,Peder Lund,James M Stafford,Benjamin A Garcia,Beatrix Ueberheide,Danny Reinberg

    The histone methyltransferase activity of PRC2 is central to the formation of H3K27me3-decorated facultative heterochromatin and gene silencing. In addition, PRC2 has been shown to automethylate its core subunits, EZH1/EZH2 and SUZ12. Here, we identify the lysine residues at which EZH1/EZH2 are automethylated with EZH2-K510 and EZH2-K514 being the major such sites in vivo. Automethylated EZH2/PRC2 exhibits a higher level of histone methyltransferase activity and is required for attaining proper cellular levels of H3K27me3. While occurring independently of PRC2 recruitment to chromatin, automethylation promotes PRC2 accessibility to the histone H3 tail. Intriguingly, EZH2 automethylation is significantly reduced in diffuse intrinsic pontine glioma (DIPG) cells that carry a lysine-to-methionine substitution in histone H3 (H3K27M), but not in cells that carry either EZH2 or EED mutants that abrogate PRC2 allosteric activation, indicating that H3K27M impairs the intrinsic activity of PRC2. Our study demonstrates a PRC2 self-regulatory mechanism through its EZH1/2-mediated automethylation activity.

    更新日期:2019-11-01
  • miR-26 suppresses adipocyte progenitor differentiation and fat production by targeting Fbxl19.
    Genes Dev. (IF 8.990) Pub Date : 2019-09-07
    Asha Acharya,Daniel C Berry,He Zhang,Yuwei Jiang,Benjamin T Jones,Robert E Hammer,Jonathan M Graff,Joshua T Mendell

    Fat storage in adult mammals is a highly regulated process that involves the mobilization of adipocyte progenitor cells (APCs) that differentiate to produce new adipocytes. Here we report a role for the broadly conserved miR-26 family of microRNAs (miR-26a-1, miR-26a-2, and miR-26b) as major regulators of APC differentiation and adipose tissue mass. Deletion of all miR-26-encoding loci in mice resulted in a dramatic expansion of adipose tissue in adult animals fed normal chow. Conversely, transgenic overexpression of miR-26a protected mice from high-fat diet-induced obesity. These effects were attributable to a cell-autonomous function of miR-26 as a potent inhibitor of APC differentiation. miR-26 blocks adipogenesis, at least in part, by repressing expression of Fbxl19, a conserved miR-26 target without a previously known role in adipocyte biology that encodes a component of SCF-type E3 ubiquitin ligase complexes. These findings have therefore revealed a novel pathway that plays a critical role in regulating adipose tissue formation in vivo and suggest new potential therapeutic targets for obesity and related disorders.

    更新日期:2019-11-01
  • Regulation of histone methylation by automethylation of PRC2.
    Genes Dev. (IF 8.990) Pub Date : 2019-09-07
    Xueyin Wang,Yicheng Long,Richard D Paucek,Anne R Gooding,Thomas Lee,Rachel M Burdorf,Thomas R Cech

    Polycomb-repressive complex 2 (PRC2) is a histone methyltransferase that is critical for regulating transcriptional repression in mammals. Its catalytic subunit, EZH2, is responsible for the trimethylation of H3K27 and also undergoes automethylation. Using mass spectrometry analysis of recombinant human PRC2, we identified three methylated lysine residues (K510, K514, and K515) on a disordered but highly conserved loop of EZH2. Methylation of these lysines increases PRC2 histone methyltransferase activity, whereas their mutation decreases activity in vitro. De novo histone methylation in an EZH2 knockout cell line is greatly impeded by mutation of the automethylation lysines. EZH2 automethylation occurs intramolecularly (in cis) by methylation of a pseudosubstrate sequence on a flexible loop. This posttranslational modification and cis regulation of PRC2 are analogous to the activation of many protein kinases by autophosphorylation. We propose that EZH2 automethylation allows PRC2 to modulate its histone methyltransferase activity by sensing histone H3 tails, SAM concentration, and perhaps other effectors.

    更新日期:2019-11-01
  • An improved auxin-inducible degron system preserves native protein levels and enables rapid and specific protein depletion.
    Genes Dev. (IF 8.990) Pub Date : 2019-08-31
    Kizhakke Mattada Sathyan,Brian D McKenna,Warren D Anderson,Fabiana M Duarte,Leighton Core,Michael J Guertin

    Rapid perturbation of protein function permits the ability to define primary molecular responses while avoiding downstream cumulative effects of protein dysregulation. The auxin-inducible degron (AID) system was developed as a tool to achieve rapid and inducible protein degradation in nonplant systems. However, tagging proteins at their endogenous loci results in chronic auxin-independent degradation by the proteasome. To correct this deficiency, we expressed the auxin response transcription factor (ARF) in an improved inducible degron system. ARF is absent from previously engineered AID systems but is a critical component of native auxin signaling. In plants, ARF directly interacts with AID in the absence of auxin, and we found that expression of the ARF PB1 (Phox and Bem1) domain suppresses constitutive degradation of AID-tagged proteins. Moreover, the rate of auxin-induced AID degradation is substantially faster in the ARF-AID system. To test the ARF-AID system in a quantitative and sensitive manner, we measured genome-wide changes in nascent transcription after rapidly depleting the ZNF143 transcription factor. Transcriptional profiling indicates that ZNF143 activates transcription in cis and regulates promoter-proximal paused RNA polymerase density. Rapidly inducible degradation systems that preserve the target protein's native expression levels and patterns will revolutionize the study of biological systems by enabling specific and temporally defined protein dysregulation.

    更新日期:2019-11-01
  • Control of homologous recombination by the HROB-MCM8-MCM9 pathway.
    Genes Dev. (IF 8.990) Pub Date : 2019-08-31
    Nicole Hustedt,Yuichiro Saito,Michal Zimmermann,Alejandro Álvarez-Quilón,Dheva Setiaputra,Salomé Adam,Andrea McEwan,Jing Yi Yuan,Michele Olivieri,Yichao Zhao,Masato T Kanemaki,Andrea Jurisicova,Daniel Durocher

    DNA repair by homologous recombination (HR) is essential for genomic integrity, tumor suppression, and the formation of gametes. HR uses DNA synthesis to repair lesions such as DNA double-strand breaks and stalled DNA replication forks, but despite having a good understanding of the steps leading to homology search and strand invasion, we know much less of the mechanisms that establish recombination-associated DNA polymerization. Here, we report that C17orf53/HROB is an OB-fold-containing factor involved in HR that acts by recruiting the MCM8-MCM9 helicase to sites of DNA damage to promote DNA synthesis. Mice with targeted mutations in Hrob are infertile due to depletion of germ cells and display phenotypes consistent with a prophase I meiotic arrest. The HROB-MCM8-MCM9 pathway acts redundantly with the HELQ helicase, and cells lacking both HROB and HELQ have severely impaired HR, suggesting that they underpin two major routes for the completion of HR downstream from RAD51. The function of HROB in HR is reminiscent of that of gp59, which acts as the replicative helicase loader during bacteriophage T4 recombination-dependent DNA replication. We therefore propose that the loading of MCM8-MCM9 by HROB may similarly be a key step in the establishment of mammalian recombination-associated DNA synthesis.

    更新日期:2019-11-01
  • Molecular basis for GIGYF-Me31B complex assembly in 4EHP-mediated translational repression.
    Genes Dev. (IF 8.990) Pub Date : 2019-08-24
    Daniel Peter,Vincenzo Ruscica,Praveen Bawankar,Ramona Weber,Sigrun Helms,Eugene Valkov,Cátia Igreja,Elisa Izaurralde

    GIGYF (Grb10-interacting GYF [glycine-tyrosine-phenylalanine domain]) proteins coordinate with 4EHP (eIF4E [eukaryotic initiation factor 4E] homologous protein), the DEAD (Asp-Glu-Ala-Asp)-box helicase Me31B/DDX6, and mRNA-binding proteins to elicit transcript-specific repression. However, the underlying molecular mechanism remains unclear. Here, we report that GIGYF contains a motif necessary and sufficient for direct interaction with Me31B/DDX6. A 2.4 Å crystal structure of the GIGYF-Me31B complex reveals that this motif arranges into a coil connected to a β hairpin on binding to conserved hydrophobic patches on the Me31B RecA2 domain. Structure-guided mutants indicate that 4EHP-GIGYF-DDX6 complex assembly is required for tristetraprolin-mediated down-regulation of an AU-rich mRNA, thus revealing the molecular principles of translational repression.

    更新日期:2019-11-01
  • A MED13-dependent skeletal muscle gene program controls systemic glucose homeostasis and hepatic metabolism.
    Genes Dev. (IF 8.990) Pub Date : 2016-02-18
    Leonela Amoasii,William Holland,Efrain Sanchez-Ortiz,Kedryn K Baskin,Mackenzie Pearson,Shawn C Burgess,Benjamin R Nelson,Rhonda Bassel-Duby,Eric N Olson

    The Mediator complex governs gene expression by linking upstream signaling pathways with the basal transcriptional machinery. However, how individual Mediator subunits may function in different tissues remains to be investigated. Through skeletal muscle-specific deletion of the Mediator subunit MED13 in mice, we discovered a gene regulatory mechanism by which skeletal muscle modulates the response of the liver to a high-fat diet. Skeletal muscle-specific deletion of MED13 in mice conferred resistance to hepatic steatosis by activating a metabolic gene program that enhances muscle glucose uptake and storage as glycogen. The consequent insulin-sensitizing effect within skeletal muscle lowered systemic glucose and insulin levels independently of weight gain and adiposity and prevented hepatic lipid accumulation. MED13 suppressed the expression of genes involved in glucose uptake and metabolism in skeletal muscle by inhibiting the nuclear receptor NURR1 and the MEF2 transcription factor. These findings reveal a fundamental molecular mechanism for the governance of glucose metabolism and the control of hepatic lipid accumulation by skeletal muscle. Intriguingly, MED13 exerts opposing metabolic actions in skeletal muscle and the heart, highlighting the customized, tissue-specific functions of the Mediator complex.

    更新日期:2019-11-01
  • Daedalus and Gasz recruit Armitage to mitochondria, bringing piRNA precursors to the biogenesis machinery.
    Genes Dev. (IF 8.990) Pub Date : 2019-05-28
    Marzia Munafò,Vera Manelli,Federica A Falconio,Ashley Sawle,Emma Kneuss,Evelyn L Eastwood,Jun Wen Eugene Seah,Benjamin Czech,Gregory J Hannon

    The Piwi-interacting RNA (piRNA) pathway is a small RNA-based immune system that silences mobile genetic elements in animal germlines. piRNA biogenesis requires a specialized machinery that converts long single-stranded precursors into small RNAs of ∼25-nucleotides in length. This process involves factors that operate in two different subcellular compartments: the nuage/Yb body and mitochondria. How these two sites communicate to achieve accurate substrate selection and efficient processing remains unclear. Here, we investigate a previously uncharacterized piRNA biogenesis factor, Daedalus (Daed), that is located on the outer mitochondrial membrane. Daed is essential for Zucchini-mediated piRNA production and the correct localization of the indispensable piRNA biogenesis factor Armitage (Armi). We found that Gasz and Daed interact with each other and likely provide a mitochondrial "anchoring platform" to ensure that Armi is held in place, proximal to Zucchini, during piRNA processing. Our data suggest that Armi initially identifies piRNA precursors in nuage/Yb bodies in a manner that depends on Piwi and then moves to mitochondria to present precursors to the mitochondrial biogenesis machinery. These results represent a significant step in understanding a critical aspect of transposon silencing; namely, how RNAs are chosen to instruct the piRNA machinery in the nature of its silencing targets.

    更新日期:2019-11-01
  • Adult and larval photoreceptors use different mechanisms to specify the same Rhodopsin fates.
    Genes Dev. (IF 8.990) Pub Date : 2007-09-06
    Simon G Sprecher,Franck Pichaud,Claude Desplan

    Although development of the adult Drosophila compound eye is very well understood, little is known about development of photoreceptors (PRs) in the simple larval eye. We show here that the larval eye is composed of 12 PRs, four of which express blue-sensitive rhodopsin5 (rh5) while the other eight contain green-sensitive rh6. This is similar to the 30:70 ratio of adult blue and green R8 cells. However, the stochastic choice of adult color PRs and the bistable loop of the warts and melted tumor suppressor genes that unambiguously specify rh5 and rh6 in R8 PRs are not involved in specification of larval PRs. Instead, primary PR precursors signal via EGFR to surrounding tissue to develop as secondary precursors, which will become Rh6-expressing PRs. EGFR signaling is required for the survival of the Rh6 subtype. Primary precursors give rise to the Rh5 subtype. Furthermore, the combinatorial action of the transcription factors Spalt, Seven-up, and Orthodenticle specifies the two PR subtypes. Therefore, even though the larval PRs and adult R8 PRs express the same rhodopsins (rh5 and rh6), they use very distinct mechanisms for their specification.

    更新日期:2019-11-01
  • The Caenorhabditis elegans gene sem-4 controls neuronal and mesodermal cell development and encodes a zinc finger protein.
    Genes Dev. (IF 8.990) Pub Date : 1996-08-01
    M Basson,H R Horvitz

    Neuronal and mesodermal cell types are generated in separate cell lineages during the larval development of Caenorhabditis elegans. Here we demonstrate that the gene sem-4 is required in both types of lineages for the normal development of neuronal and mesodermal cell types. The sem-4 gene encodes a protein containing seven zinc finger motifs of the C2H2 class, four of which are arranged in two pairs widely separated in the primary sequence of the protein. These pairs of zinc fingers are similar to pairs of zinc fingers in the protein encoded by the Drosophila homeotic gene spalt and in the human transcription factor PRDII-BF1. Analysis of sem-4 alleles suggests that different zinc fingers in the SEM-4 protein may function differentially in neuronal and mesodermal cell types. We propose that sem-4 interacts with different transcription factors in different cell types to control the transcription of genes that function in the processes of neuronal and mesodermal cell development.

    更新日期:2019-11-01
  • CDK12 phosphorylates 4E-BP1 to enable mTORC1-dependent translation and mitotic genome stability.
    Genes Dev. (IF 8.990) Pub Date : 2019-03-02
    Seung H Choi,Thomas F Martinez,Seongjae Kim,Cynthia Donaldson,Maxim N Shokhirev,Alan Saghatelian,Katherine A Jones

    The RNA polymerase II (RNAPII) C-terminal domain kinase, CDK12, regulates genome stability, expression of DNA repair genes, and cancer cell resistance to chemotherapy and immunotherapy. In addition to its role in mRNA biosynthesis of DNA repair genes, we show here that CDK12 phosphorylates the mRNA 5' cap-binding repressor, 4E-BP1, to promote translation of mTORC1-dependent mRNAs. In particular, we found that phosphorylation of 4E-BP1 by mTORC1 (T37 and T46) facilitates subsequent CDK12 phosphorylation at two Ser-Pro sites (S65 and T70) that control the exchange of 4E-BP1 with eIF4G at the 5' cap of CHK1 and other target mRNAs. RNA immunoprecipitation coupled with deep sequencing (RIP-seq) revealed that CDK12 regulates release of 4E-BP1, and binding of eIF4G, to many mTORC1 target mRNAs, including those needed for MYC transformation. Genome-wide ribosome profiling (Ribo-seq) further identified specific CDK12 "translation-only" target mRNAs, including many mTORC1 target mRNAs as well as many subunits of mitotic and centromere/centrosome complexes. Accordingly, confocal imaging analyses revealed severe chromosome misalignment, bridging, and segregation defects in cells deprived of CDK12 or CCNK. We conclude that the nuclear RNAPII-CTD kinase CDK12 cooperates with mTORC1, and controls a specialized translation network that is essential for mitotic chromosome stability.

    更新日期:2019-11-01
  • High telomerase is a hallmark of undifferentiated spermatogonia and is required for maintenance of male germline stem cells.
    Genes Dev. (IF 8.990) Pub Date : 2015-11-21
    Matthew F Pech,Alina Garbuzov,Kazuteru Hasegawa,Meena Sukhwani,Ruixuan J Zhang,Bérénice A Benayoun,Stephanie A Brockman,Shengda Lin,Anne Brunet,Kyle E Orwig,Steven E Artandi

    Telomerase inactivation causes loss of the male germline in worms, fish, and mice, indicating a conserved dependence on telomere maintenance in this cell lineage. Here, using telomerase reverse transcriptase (Tert) reporter mice, we found that very high telomerase expression is a hallmark of undifferentiated spermatogonia, the mitotic population where germline stem cells reside. We exploited these high telomerase levels as a basis for purifying undifferentiated spermatogonia using fluorescence-activated cell sorting. Telomerase levels in undifferentiated spermatogonia and embryonic stem cells are comparable and much greater than in somatic progenitor compartments. Within the germline, we uncovered an unanticipated gradient of telomerase activity that also enables isolation of more mature populations. Transcriptomic comparisons of Tert(High) undifferentiated spermatogonia and Tert(Low) differentiated spermatogonia by RNA sequencing reveals marked differences in cell cycle and key molecular features of each compartment. Transplantation studies show that germline stem cell activity is confined to the Tert(High) cKit(-) population. Telomere shortening in telomerase knockout strains causes depletion of undifferentiated spermatogonia and eventual loss of all germ cells after undifferentiated spermatogonia drop below a critical threshold. These data reveal that high telomerase expression is a fundamental characteristic of germline stem cells, thus explaining the broad dependence on telomerase for germline immortality in metazoans.

    更新日期:2019-11-01
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