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  • Sec17 (α-SNAP) and an SM-tethering complex regulate the outcome of SNARE zippering in vitro and in vivo
    eLife (IF 7.725) Pub Date : 2017-09-19
    Matthew L Schwartz, Daniel P Nickerson, Braden T Lobingier, Rachael L Plemel, Mengtong Duan, Cortney G Angers, Michael Zick, Alexey J Merz

    Zippering of SNARE complexes spanning docked membranes is essential for most intracellular fusion events. Here we explore how SNARE regulators operate on discrete zippering states. The formation of a metastable trans-complex, catalyzed by HOPS and its SM subunit Vps33, is followed by subsequent zippering transitions that increase the probability of fusion. Operating independently of Sec18 (NSF) catalysis, Sec17 (α-SNAP) either inhibits or stimulates SNARE-mediated fusion. If HOPS or Vps33 are absent, Sec17 inhibits fusion at an early stage. Thus, Vps33/HOPS promotes productive SNARE assembly in the presence of otherwise inhibitory Sec17. Once SNAREs are partially zipped, Sec17 promotes fusion in either the presence or absence of HOPS, but with faster kinetics when HOPS is absent, suggesting that ejection of the SM is a rate-limiting step.

    更新日期:2017-09-19
  • The C. elegans neural editome reveals an ADAR target mRNA required for proper chemotaxis
    eLife (IF 7.725) Pub Date : 2017-09-19
    Sarah N Deffit, Brian A Yee, Aidan C Manning, Suba Rajendren, Pranathi Vadlamani, Emily C Wheeler, Alain Domissy, Michael C Washburn, Gene W Yeo, Heather A Hundley

    ADAR proteins alter gene expression both by catalyzing adenosine (A) to inosine (I) RNA editing and binding to regulatory elements in target RNAs. Loss of ADARs affects neuronal function in all animals studied to date. Caenorhabditis elegans lacking ADARs exhibit reduced chemotaxis, but the targets responsible for this phenotype remain unknown. To identify critical neural ADAR targets in C. elegans, we performed an unbiased assessment of the effects of ADR-2, the only A-to-I editing enzyme in C. elegans, on the neural transcriptome. Development and implementation of publicly available software, SAILOR, identified 7,361 A-to-I editing events across the neural transcriptome. Intersecting the neural editome with adr-2 associated gene expression changes, revealed an edited mRNA, clec-41, whose neural expression is dependent on deamination. Restoring clec-41 expression in adr-2 deficient neural cells rescued the chemotaxis defect, providing the first evidence that neuronal phenotypes of ADAR mutants can be caused by altered gene expression.

    更新日期:2017-09-19
  • Computational design of environmental sensors for the potent opioid fentanyl
    eLife (IF 7.725) Pub Date : 2017-09-19
    Matthew J Bick, Per J Greisen, Kevin J Morey, Mauricio S Antunes, David La, Banumathi Sankaran, Luc Reymond, Kai Johnsson, June I Medford, David Baker

    We describe the computational design of proteins that bind the potent analgesic fentanyl. Our approach employs a fast docking algorithm to find shape complementary ligand placement in protein scaffolds, followed by design of the surrounding residues to optimize binding affinity. Co-crystal structures of the highest affinity binder reveal a highly preorganized binding site, and an overall architecture and ligand placement in close agreement with the design model. We use the designs to generate plant sensors for fentanyl by coupling ligand binding to design stability. The method should be generally useful for detecting toxic hydrophobic compounds in the environment.

    更新日期:2017-09-19
  • Preconditioned cues have no value
    eLife (IF 7.725) Pub Date : 2017-09-19
    Melissa Sharpe, Hannah Batchelor, Geoffrey Schoenbaum

    Sensory preconditioning has been used to implicate midbrain dopamine in model-based learning, contradicting the view that dopamine transients reflect model-free value. However, it has been suggested that model-free value might accrue directly to the preconditioned cue through mediated learning. Here, building on previous work (Sadacca et al., 2016), we address this question by testing whether a preconditioned cue will support conditioned reinforcement in rats. We found that while both directly conditioned and second-order conditioned cues supported robust conditioned reinforcement, a preconditioned cue did not. These data show that the preconditioned cue in our procedure does not directly accrue model-free value and further suggest that the cue may not necessarily access value even indirectly in a model-based manner. If so, then phasic response of dopamine neurons to cues in this setting cannot be described as signaling errors in predicting value.

    更新日期:2017-09-19
  • Loss of foxo rescues stem cell aging in Drosophila germ line
    eLife (IF 7.725) Pub Date : 2017-09-19
    Filippo Artoni, Rebecca Kreipke, Ondina Palmeira, Connor Dixon, Zachary Goldberg, Hannele Ruohola-Baker

    Aging stem cells lose the capacity to properly respond to injury and regenerate their residing tissues. Here, we utilized the ability of Drosophila melanogaster germline stem cells (GSCs) to survive exposure to low doses of ionizing radiation (IR) as a model of adult stem cell injury and identified a regeneration defect in aging GSCs: while aging GSCs survive exposure to IR, they fail to reenter the cell cycle and regenerate the germline in a timely manner. Mechanistically, we identify foxo and mTOR homologue, Tor as important regulators of GSC quiescence following exposure to ionizing radiation. foxo is required for entry in quiescence, while Tor is essential for cell cycle reentry. Importantly, we further show that the lack of regeneration in aging germ line stem cells after IR can be rescued by loss of foxo.

    更新日期:2017-09-19
  • Major transcriptional changes observed in the Fulani, an ethnic group less susceptible to malaria
    eLife (IF 7.725) Pub Date : 2017-09-19
    Jaclyn E Quin, Ioana Bujila, Mariama Chérif, Guillaume S Sanou, Ying Qu, Manijeh Vafa Homann, Anna Rolicka, Sodiomon B Sirima, Mary A O'Connell, Andreas Lennartsson, Marita Troye-Blomberg, Issa Nebie, Ann-Kristin Östlund Farrants

    The Fulani ethnic group has relatively better protection from Plasmodium falciparum malaria, as reflected by fewer symptomatic cases of malaria, lower infection rates, and lower parasite densities compared to sympatric ethnic groups<strong>.</strong> However, the basis for this lower susceptibility to malaria by the Fulani is unknown. We have performed a pilot study to examine global transcription and DNA methylation patterns in specific immune cell populations in the Fulani to elucidate the mechanisms that confer the lower susceptibility to P.falciparum malaria. When we compared uninfected and infected Fulani individuals, in contrast to uninfected and infected individuals from the sympatric ethnic group Mossi, we observed a key difference: a strong transcriptional response was only detected in the monocyte fraction of the Fulani, where over 1000 genes were significantly differentially expressed upon P.falciparum infection.

    更新日期:2017-09-19
  • Computational design of environmental sensors for the potent opioid fentanyl
    eLife (IF 7.725) Pub Date : 2017-09-19
    Matthew J Bick, Per J Greisen, Kevin J Morey, Mauricio S Antunes, David La, Banumathi Sankaran, Luc Reymond, Kai Johnsson, June I Medford, David Baker

    We describe the computational design of proteins that bind the potent analgesic fentanyl. Our approach employs a fast docking algorithm to find shape complementary ligand placement in protein scaffolds, followed by design of the surrounding residues to optimize binding affinity. Co-crystal structures of the highest affinity binder reveal a highly preorganized binding site, and an overall architecture and ligand placement in close agreement with the design model. We use the designs to generate plant sensors for fentanyl by coupling ligand binding to design stability. The method should be generally useful for detecting toxic hydrophobic compounds in the environment.

    更新日期:2017-09-19
  • MELK is not necessary for the proliferation of basal-like breast cancer cells
    eLife (IF 7.725) Pub Date : 2017-09-19
    Hai-Tsang Huang, Hyuk-Soo Seo, Tinghu Zhang, Yubao Wang, Baishan Jiang, Qing Li, Dennis L Buckley, Behnam Nabet, Justin M Roberts, Joshiawa Paulk, Shiva Dastjerdi, Georg E Winter, Hilary McLauchlan, Jennifer Moran, James E Bradner, Michael J Eck, Sirano Dhe-Paganon, Jean J Zhao, Nathanael S Gray

    Thorough preclinical target validation is essential for the success of drug discovery efforts. In this study, we combined chemical and genetic perturbants, including the development of a novel selective maternal embryonic leucine zipper kinase (MELK) inhibitor HTH-01-091, CRISPR/Cas9-mediated MELK knockout, a novel chemical-induced protein degradation strategy, RNA interference and CRISPR interference to validate MELK as a therapeutic target in basal-like breast cancers (BBC). In common culture conditions, we found that small molecule inhibition, genetic deletion, or acute depletion of MELK did not significantly affect cellular growth. This discrepancy to previous findings illuminated selectivity issues of the widely used MELK inhibitor OTSSP167, and potential off-target effects of MELK-targeting short hairpins. The different genetic and chemical tools developed here allow for the identification and validation of any causal roles MELK may play in cancer biology, which will be required to guide future MELK drug discovery efforts. Furthermore, our study provides a general framework for preclinical target validation.https://doi.org/10.7554/eLife.26693.001

    更新日期:2017-09-19
  • Loss of foxo rescues stem cell aging in Drosophila germ line
    eLife (IF 7.725) Pub Date : 2017-09-19
    Filippo Artoni, Rebecca Kreipke, Ondina Palmeira, Connor Dixon, Zachary Goldberg, Hannele Ruohola-Baker

    Aging stem cells lose the capacity to properly respond to injury and regenerate their residing tissues. Here, we utilized the ability of Drosophila melanogaster germline stem cells (GSCs) to survive exposure to low doses of ionizing radiation (IR) as a model of adult stem cell injury and identified a regeneration defect in aging GSCs: while aging GSCs survive exposure to IR, they fail to reenter the cell cycle and regenerate the germline in a timely manner. Mechanistically, we identify foxo and mTOR homologue, Tor as important regulators of GSC quiescence following exposure to ionizing radiation. foxo is required for entry in quiescence, while Tor is essential for cell cycle reentry. Importantly, we further show that the lack of regeneration in aging germ line stem cells after IR can be rescued by loss of foxo.

    更新日期:2017-09-19
  • Preconditioned cues have no value
    eLife (IF 7.725) Pub Date : 2017-09-19
    Melissa Sharpe, Hannah Batchelor, Geoffrey Schoenbaum

    Sensory preconditioning has been used to implicate midbrain dopamine in model-based learning, contradicting the view that dopamine transients reflect model-free value. However, it has been suggested that model-free value might accrue directly to the preconditioned cue through mediated learning. Here, building on previous work (Sadacca et al., 2016), we address this question by testing whether a preconditioned cue will support conditioned reinforcement in rats. We found that while both directly conditioned and second-order conditioned cues supported robust conditioned reinforcement, a preconditioned cue did not. These data show that the preconditioned cue in our procedure does not directly accrue model-free value and further suggest that the cue may not necessarily access value even indirectly in a model-based manner. If so, then phasic response of dopamine neurons to cues in this setting cannot be described as signaling errors in predicting value.

    更新日期:2017-09-19
  • BMPs direct sensory interneuron identity in the developing spinal cord using signal-specific not morphogenic activities
    eLife (IF 7.725) Pub Date : 2017-09-19
    Madeline G Andrews, Lorenzo M del Castillo, Eliana Ochoa-Bolton, Ken Yamauchi, Jan Smogorzewski, Samantha J Butler

    The Bone Morphogenetic Protein (BMP) family reiteratively signals to direct disparate cellular fates throughout embryogenesis. In the developing dorsal spinal cord, multiple BMPs are required to specify sensory interneurons (INs). Previous studies suggested that the BMPs act as concentration-dependent morphogens to direct IN identity, analogous to the manner in which sonic hedgehog patterns the ventral spinal cord. However, it remains unresolved how multiple BMPs would cooperate to establish a unified morphogen gradient. Our studies support an alternative model: BMPs have signal-specific activities directing particular IN fates. Using chicken and mouse models, we show that the identity, not concentration, of the BMP ligand directs distinct dorsal identities. Individual BMPs promote progenitor patterning or neuronal differentiation by their activation of different type I BMP receptors and distinct modulations of the cell cycle. Together, this study shows that a ‘mix and match’ code of BMP signaling results in distinct classes of sensory INs.https://doi.org/10.7554/eLife.30647.001

    更新日期:2017-09-19
  • Restraint of presynaptic protein levels by Wnd/DLK signaling mediates synaptic defects associated with the kinesin-3 motor Unc-104
    eLife (IF 7.725) Pub Date : 2017-09-19
    Jiaxing Li, Yao V Zhang, Elham Asghari Adib, Doychin T Stanchev, Xin Xiong, Susan Klinedinst, Pushpanjali Soppina, Thomas Robert Jahn, Richard I Hume, Tobias M Rasse, Catherine A Collins

    The kinesin-3 family member Unc-104/KIF1A is required for axonal transport of many presynaptic components to synapses, and mutation of this gene results in synaptic dysfunction in mice, flies and worms. Our studies at the Drosophila neuromuscular junction indicate that many synaptic defects in unc-104-null mutants are mediated independently of Unc-104’s transport function, via the Wallenda (Wnd)/DLK MAP kinase axonal damage signaling pathway. Wnd signaling becomes activated when Unc-104’s function is disrupted, and leads to impairment of synaptic structure and function by restraining the expression level of active zone (AZ) and synaptic vesicle (SV) components. This action concomitantly suppresses the buildup of synaptic proteins in neuronal cell bodies, hence may play an adaptive role to stresses that impair axonal transport. Wnd signaling also becomes activated when pre-synaptic proteins are over-expressed, suggesting the existence of a feedback circuit to match synaptic protein levels to the transport capacity of the axon.https://doi.org/10.7554/eLife.24271.001

    更新日期:2017-09-19
  • Cyclic bouts of extreme bradycardia counteract the high metabolism of frugivorous bats
    eLife (IF 7.725) Pub Date : 2017-09-19
    M Teague O'Mara, Martin Wikelski, Christian C Voigt, Andries Ter Maat, Henry S Pollock, Gary Burness, Lanna M Desantis, Dina KN Dechmann

    Active flight requires the ability to efficiently fuel bursts of costly locomotion while maximizing energy conservation during non-flying times. We took a multi-faceted approach to estimate how fruit-eating bats (Uroderma bilobatum) manage a high-energy lifestyle fueled primarily by fig juice. Miniaturized heart rate telemetry shows that they use a novel, cyclic, bradycardic state that reduces daily energetic expenditure by 10% and counteracts heart rates as high as 900 bpm during flight. Uroderma bilobatum support flight with some of the fastest metabolic incorporation rates and dynamic circulating cortisol in vertebrates. These bats will exchange fat reserves within 24 hr, meaning that they must survive on the food of the day and are at daily risk of starvation. Energetic flexibly in U. bilobatum highlights the fundamental role of ecological pressures on integrative energetic networks and the still poorly understood energetic strategies of animals in the tropics.https://doi.org/10.7554/eLife.26686.001

    更新日期:2017-09-19
  • Biofilms: Flipping the switch
    eLife (IF 7.725) Pub Date : 2017-09-19
    Xavier Pierrat, Alexandre Persat

    A structural switch controls the architecture of Vibrio cholerae biofilms by mediating the interactions between two matrix components.

    更新日期:2017-09-19
  • Major transcriptional changes observed in the Fulani, an ethnic group less susceptible to malaria
    eLife (IF 7.725) Pub Date : 2017-09-19
    Jaclyn E Quin, Ioana Bujila, Mariama Chérif, Guillaume S Sanou, Ying Qu, Manijeh Vafa Homann, Anna Rolicka, Sodiomon B Sirima, Mary A O'Connell, Andreas Lennartsson, Marita Troye-Blomberg, Issa Nebie, Ann-Kristin Östlund Farrants

    The Fulani ethnic group has relatively better protection from Plasmodium falciparum malaria, as reflected by fewer symptomatic cases of malaria, lower infection rates, and lower parasite densities compared to sympatric ethnic groups<strong>.</strong> However, the basis for this lower susceptibility to malaria by the Fulani is unknown. We have performed a pilot study to examine global transcription and DNA methylation patterns in specific immune cell populations in the Fulani to elucidate the mechanisms that confer the lower susceptibility to P.falciparum malaria. When we compared uninfected and infected Fulani individuals, in contrast to uninfected and infected individuals from the sympatric ethnic group Mossi, we observed a key difference: a strong transcriptional response was only detected in the monocyte fraction of the Fulani, where over 1000 genes were significantly differentially expressed upon P.falciparum infection.

    更新日期:2017-09-19
  • The C. elegans neural editome reveals an ADAR target mRNA required for proper chemotaxis
    eLife (IF 7.725) Pub Date : 2017-09-19
    Sarah N Deffit, Brian A Yee, Aidan C Manning, Suba Rajendren, Pranathi Vadlamani, Emily C Wheeler, Alain Domissy, Michael C Washburn, Gene W Yeo, Heather A Hundley

    ADAR proteins alter gene expression both by catalyzing adenosine (A) to inosine (I) RNA editing and binding to regulatory elements in target RNAs. Loss of ADARs affects neuronal function in all animals studied to date. Caenorhabditis elegans lacking ADARs exhibit reduced chemotaxis, but the targets responsible for this phenotype remain unknown. To identify critical neural ADAR targets in C. elegans, we performed an unbiased assessment of the effects of ADR-2, the only A-to-I editing enzyme in C. elegans, on the neural transcriptome. Development and implementation of publicly available software, SAILOR, identified 7,361 A-to-I editing events across the neural transcriptome. Intersecting the neural editome with adr-2 associated gene expression changes, revealed an edited mRNA, clec-41, whose neural expression is dependent on deamination. Restoring clec-41 expression in adr-2 deficient neural cells rescued the chemotaxis defect, providing the first evidence that neuronal phenotypes of ADAR mutants can be caused by altered gene expression.

    更新日期:2017-09-19
  • Sec17 (α-SNAP) and an SM-tethering complex regulate the outcome of SNARE zippering in vitro and in vivo
    eLife (IF 7.725) Pub Date : 2017-09-19
    Matthew L Schwartz, Daniel P Nickerson, Braden T Lobingier, Rachael L Plemel, Mengtong Duan, Cortney G Angers, Michael Zick, Alexey J Merz

    Zippering of SNARE complexes spanning docked membranes is essential for most intracellular fusion events. Here we explore how SNARE regulators operate on discrete zippering states. The formation of a metastable trans-complex, catalyzed by HOPS and its SM subunit Vps33, is followed by subsequent zippering transitions that increase the probability of fusion. Operating independently of Sec18 (NSF) catalysis, Sec17 (α-SNAP) either inhibits or stimulates SNARE-mediated fusion. If HOPS or Vps33 are absent, Sec17 inhibits fusion at an early stage. Thus, Vps33/HOPS promotes productive SNARE assembly in the presence of otherwise inhibitory Sec17. Once SNAREs are partially zipped, Sec17 promotes fusion in either the presence or absence of HOPS, but with faster kinetics when HOPS is absent, suggesting that ejection of the SM is a rate-limiting step.

    更新日期:2017-09-19
  • Dynamic representation of partially occluded objects in primate prefrontal and visual cortex
    eLife (IF 7.725) Pub Date : 2017-09-19
    Amber M Fyall, Yasmine El-Shamayleh, Hannah Choi, Eric Shea-Brown, Anitha Pasupathy

    Successful recognition of partially occluded objects is presumed to involve dynamic interactions between brain areas responsible for vision and cognition, but neurophysiological evidence for the involvement of feedback signals is lacking. Here, we demonstrate that neurons in the ventrolateral prefrontal cortex (vlPFC) of monkeys performing a shape discrimination task respond more strongly to occluded than unoccluded stimuli. In contrast, neurons in visual area V4 respond more strongly to unoccluded stimuli. Analyses of V4 response dynamics reveal that many neurons exhibit two transient response peaks, the second of which emerges after vlPFC response onset and displays stronger selectivity for occluded shapes. We replicate these findings using a model of V4/vlPFC interactions in which occlusion-sensitive vlPFC neurons feed back to shape-selective V4 neurons, thereby enhancing V4 responses and selectivity to occluded shapes. These results reveal how signals from frontal and visual cortex could interact to facilitate object recognition under occlusion.https://doi.org/10.7554/eLife.25784.001

    更新日期:2017-09-19
  • Protein-mediated RNA folding governs sequence-specific interactions between rotavirus genome segments
    eLife (IF 7.725) Pub Date : 2017-09-18
    Alexander Borodavka, Eric C Dykeman, Waldemar Schrimpf, Don C Lamb

    Segmented RNA viruses are ubiquitous pathogens, which include influenza viruses and rotaviruses. A major challenge in understanding their assembly is the combinatorial problem of a non-random selection of a full genomic set of distinct RNAs. This process involves complex RNA-RNA and protein-RNA interactions, which are often obscured by non-specific binding at concentrations approaching in vivo assembly conditions. Here, we present direct experimental evidence of sequence-specific inter-segment interactions between rotavirus RNAs, taking place in a complex RNA- and protein-rich milieu. We show that binding of the rotavirus-encoded non-structural protein NSP2 to viral ssRNAs results in the remodeling of RNA, which is conducive to formation of stable inter-segment contacts. To identify the sites of these interactions, we have developed an RNA-RNA SELEX approach for mapping the sequences involved in inter-segment base-pairing. Our findings elucidate the molecular basis underlying inter-segment interactions in rotaviruses, paving the way for delineating similar RNA-RNA interactions that govern assembly of other segmented RNA viruses. https://doi.org/10.7554/eLife.27453.001

    更新日期:2017-09-19
  • The DREAM complex through its subunit Lin37 cooperates with Rb to initiate quiescence
    eLife (IF 7.725) Pub Date : 2017-09-18
    Christina FS Mages, Axel Wintsche, Stephan H Bernhart, Gerd A Müller

    The retinoblastoma Rb protein is an important factor controlling the cell cycle. Yet, mammalian cells carrying Rb deletions are still able to arrest under growth-limiting conditions. The Rb-related proteins p107 and p130, which are components of the DREAM complex, had been suggested to be responsible for a continued ability to arrest by inhibiting E2f activity and by recruiting chromatin-modifying enzymes. Here, we show that p130 and p107 are not sufficient for DREAM-dependent repression. We identify the MuvB protein Lin37 as an essential factor for DREAM function. Cells not expressing Lin37 proliferate normally, but DREAM completely loses its ability to repress genes in G0/G1 while all remaining subunits, including p130/p107, still bind to target gene promoters. Furthermore, cells lacking both Rb and Lin37 are incapable of exiting the cell cycle. Thus, Lin37 is an essential component of DREAM that cooperates with Rb to induce quiescence. https://doi.org/10.7554/eLife.26876.001

    更新日期:2017-09-18
  • The P2X7 receptor forms a dye-permeable pore independent of its intracellular domain but dependent on membrane lipid composition
    eLife (IF 7.725) Pub Date : 2017-09-18
    Akira Karasawa, Kevin Michalski, Polina Mikhelzon, Toshimitsu Kawate

    The P2X7 receptor mediates extracellular-ATP signaling implicated in the development of devastating diseases such as chronic pain and cancer. Activation of the P2X7 receptor leads to opening of the characteristic dye-permeable membrane pore for molecules up to ~900 Da. However, it remains controversial what constitutes this peculiar pore and how it opens. Here we show that the panda P2X7 receptor, when purified and reconstituted into liposomes, forms an intrinsic dye-permeable pore in the absence of other cellular components. Unexpectedly, we found that this pore opens independent of its unique C-terminal domain. We also found that P2X7 channel activity is facilitated by phosphatidylglycerol and sphingomyelin, but dominantly inhibited by cholesterol through direct interactions with the transmembrane domain. In combination with cell-based functional studies, our data suggest that the P2X7 receptor itself constitutes a lipid-composition dependent dye-permeable pore, whose opening is facilitated by palmitoylated cysteines near the pore-lining helix.

    更新日期:2017-09-18
  • The P2X7 receptor forms a dye-permeable pore independent of its intracellular domain but dependent on membrane lipid composition
    eLife (IF 7.725) Pub Date : 2017-09-18
    Akira Karasawa, Kevin Michalski, Polina Mikhelzon, Toshimitsu Kawate

    The P2X7 receptor mediates extracellular-ATP signaling implicated in the development of devastating diseases such as chronic pain and cancer. Activation of the P2X7 receptor leads to opening of the characteristic dye-permeable membrane pore for molecules up to ~900 Da. However, it remains controversial what constitutes this peculiar pore and how it opens. Here we show that the panda P2X7 receptor, when purified and reconstituted into liposomes, forms an intrinsic dye-permeable pore in the absence of other cellular components. Unexpectedly, we found that this pore opens independent of its unique C-terminal domain. We also found that P2X7 channel activity is facilitated by phosphatidylglycerol and sphingomyelin, but dominantly inhibited by cholesterol through direct interactions with the transmembrane domain. In combination with cell-based functional studies, our data suggest that the P2X7 receptor itself constitutes a lipid-composition dependent dye-permeable pore, whose opening is facilitated by palmitoylated cysteines near the pore-lining helix.

    更新日期:2017-09-18
  • Cortex-wide BOLD fMRI activity reflects locally-recorded slow oscillation-associated calcium waves
    eLife (IF 7.725) Pub Date : 2017-09-15
    Miriam Schwalm, Florian Schmid, Lydia Wachsmuth, Hendrik Backhaus, Andrea Kronfeld, Felipe Aedo Jury, Pierre-Hugues Prouvot, Consuelo Fois, Franziska Albers, Timo van Alst, Cornelius Faber, Albrecht Stroh

    Spontaneous slow oscillation-associated slow wave activity represents an internally generated state which is characterized by alternations of network quiescence and stereotypical episodes of neuronal activity - slow wave events. However, it remains unclear which macroscopic signal is related to these active periods of the slow wave rhythm. We used optic fiber-based calcium recordings of local neural populations in cortex and thalamus to detect neurophysiologically defined slow calcium waves in isoflurane anesthetized rats. The individual slow wave events were used for an event-related analysis of simultaneously acquired whole-brain BOLD fMRI. We identified BOLD responses directly related to onsets of slow calcium waves, revealing a cortex-wide BOLD correlate: the entire cortex was engaged in this specific type of slow wave activity. These findings demonstrate a direct relation of defined neurophysiological events to a specific BOLD activity pattern and were confirmed for ongoing slow wave activity by independent component and seed-based analyses.

    更新日期:2017-09-18
  • Nanoscale architecture of the Schizosaccharomyces pombe contractile ring
    eLife (IF 7.725) Pub Date : 2017-09-15
    Nathan A McDonald, Abigail L Lind, Sarah E Smith, Rong Li, Kathleen Gould

    The contractile ring is a complex molecular apparatus important for dividing many eukaryotic cells. Despite knowledge of its composition, the molecular architecture of the ring is not known. Here we applied super-resolution microscopy and FRET to determine the nanoscale spatial organization of Schizosaccharomyces pombe contractile ring components relative to the plasma membrane. As in other membrane-tethered actin structures, contractile ring proteins are stratified relative to the membrane. The lowest layer (0-80 nm) contains membrane-binding scaffolds, formin, and the myosin-II tail. An intermediate zone (80-160 nm) consists of a network of cytokinesis accessory proteins and signaling components that influence cell division. Most interior from the membrane (160-400 nm) is F-actin, myosin motor domains, and an F-actin crosslinker. Circumferentially within the ring, multiple proximal membrane proteins form different sized clusters, while components farther from the membrane are uniformly distributed. This comprehensive organizational map provides a framework for understanding contractile ring function.

    更新日期:2017-09-18
  • Scc2/Nipbl hops between chromosomal cohesin rings after loading
    eLife (IF 7.725) Pub Date : 2017-09-15
    James DP Rhodes, Davide Mazza, Kim A Nasmyth, Stephan Uphoff

    The cohesin complex mediates DNA-DNA interactions both between (sister chromatid cohesion) and within chromosomes (DNA looping). It has been suggested that intra-chromosome loops are generated by extrusion of DNAs through the lumen of cohesin's ring. Scc2 (Nipbl) stimulates cohesin's ABC-like ATPase and is essential for loading cohesin onto chromosomes. However, it is possible that the stimulation of cohesin's ATPase by Scc2 also has a post-loading function, for example driving loop extrusion. Using fluorescence recovery after photobleaching (FRAP) and single- molecule tracking, we show that Scc2 binds dynamically to chromatin, principally through an association with cohesin. Scc2's movement within chromatin is consistent with a 'stop-and-go' or 'hopping' motion. We suggest that a low diffusion coefficient, a low stoichiometry relative to cohesin, and a high affinity for chromosomal cohesin enables Scc2 to move rapidly from one chromosomal cohesin complex to another, performing a function distinct from loading.

    更新日期:2017-09-18
  • Synaptic up-scaling preserves motor circuit output after chronic, natural inactivity
    eLife (IF 7.725) Pub Date : 2017-09-15
    Joseph M Santin, Mauricio Vallejo, Lynn K Hartzler

    Neural systems use homeostatic plasticity to maintain normal brain functions and to prevent abnormal activity. Surprisingly, homeostatic mechanisms that regulate circuit output have mainly been demonstrated during artificial and/or pathological perturbations. Natural, physiological scenarios that activate these stabilizing mechanisms in neural networks of mature animals remain elusive. To establish the extent to which a naturally inactive circuit engages mechanisms of homeostatic plasticity, we utilized the respiratory motor circuit in bullfrogs that normally remains inactive for several months during the winter. We found that inactive respiratory motoneurons exhibit a classic form of homeostatic plasticity, up-scaling of AMPA-glutamate receptors. Up-scaling increased the synaptic strength of respiratory motoneurons and acted to boost motor amplitude from the respiratory network following months of inactivity. Our results show that synaptic scaling sustains strength of the respiratory motor output following months of inactivity, thereby supporting a major neuroscience hypothesis in a normal context for an adult animal.

    更新日期:2017-09-18
  • A causal role for right frontopolar cortex in directed, but not random, exploration
    eLife (IF 7.725) Pub Date : 2017-09-15
    Wojciech K Zajkowski, Malgorzata Kossut, Robert C Wilson

    The explore-exploit dilemma occurs anytime we must choose between exploring unknown options for information and exploiting known resources for reward. Previous work suggests that people use two different strategies to solve the explore-exploit dilemma: directed exploration, driven by information seeking, and random exploration, driven by decision noise. Here, we show that these two strategies rely on different neural systems. Using transcranial magnetic stimulation to inhibit the right frontopolar cortex, we were able to selectively inhibit directed exploration while leaving random exploration intact. This suggests a causal role for right frontopolar cortex in directed, but not random, exploration and that directed and random exploration rely on (at least partially) dissociable neural systems.

    更新日期:2017-09-18
  • Lipids and ions traverse the membrane by the same physical pathway in the nhTMEM16 scramblase
    eLife (IF 7.725) Pub Date : 2017-09-16
    Tao Jiang, Kuai Yu, H Criss Hartzell, Emad Tajkhorshid

    From bacteria to mammals, different phospholipid species are segregated between the inner and outer leaflets of the plasma membrane by ATP-dependent lipid transporters. Disruption of this asymmetry by ATP-independent phospholipid scrambling is important in cellular signaling, but its mechanism remains incompletely understood. Using MD simulations coupled with experimental assays, we show that the surface hydrophilic transmembrane cavity exposed to the lipid bilayer on the fungal scramblase nhTMEM16 serves as the pathway for both lipid translocation and ion conduction across the membrane. Ca2+ binding stimulates its open conformation by altering the structure of transmembrane helices that line the cavity. We have identified key amino acids necessary for phospholipid scrambling and validated the idea that ions permeate TMEM16 Cl- channels via a structurally homologous pathway by showing that mutation of two residues in the pore region of the TMEM16A Ca2+-activated Cl- channel convert it into a robust scramblase.

    更新日期:2017-09-18
  • Lineage tracing of genome-edited alleles reveals high fidelity axolotl limb regeneration
    eLife (IF 7.725) Pub Date : 2017-09-16
    Grant Parker Flowers, Lucas D Sanor, Craig M Crews

    Salamanders are unparalleled among tetrapods in their ability to regenerate many structures, including entire limbs, and the study of this ability may provide insights into human regenerative therapies. The complex structure of the limb poses challenges to the investigation of the cellular and molecular basis of its regeneration. Using CRISPR/Cas, we genetically labelled unique cell lineages within the developing axolotl embryo and tracked the frequency of each lineage within amputated and fully regenerated limbs. This allowed us, for the first time, to assess the contributions of multiple low frequency cell lineages to the regenerating limb at once. Our comparisons reveal that regenerated limbs are high fidelity replicas of the originals even after repeated amputations.

    更新日期:2017-09-18
  • Direct modulation of aberrant brain network connectivity through real-time neurofeedback
    eLife (IF 7.725) Pub Date : 2017-09-16
    Michal Ramot, Sara Kimmich, Javier Gonzalez-Castillo, Vinai Roopchansingh, Haroon Popal, Emily White, Stephen J Gotts, Alex Martin

    The existence of abnormal connectivity patterns between resting state networks in neuropsychiatric disorders, including Autism Spectrum Disorder (ASD), has been well established. Traditional treatment methods in ASD are limited, and do not address the aberrant network structure. Using real-time fMRI neurofeedback, we directly trained 3 brain nodes in participants with ASD, in which the aberrant connectivity has been shown to correlate with symptom severity. Desired network connectivity patterns were reinforced in real-time, without participants' awareness of the training taking place. This training regimen produced large, significant long-term changes in correlations at the network level, and whole brain analysis revealed that the greatest changes were focused on the areas being trained. These changes were not found in the control group. Moreover, changes in ASD resting state connectivity following the training were correlated to changes in behavior, suggesting that neurofeedback can be used to directly alter complex, clinically relevant network connectivity patterns.

    更新日期:2017-09-18
  • Nanoscale architecture of the Schizosaccharomyces pombe contractile ring
    eLife (IF 7.725) Pub Date : 2017-09-15
    Nathan A McDonald, Abigail L Lind, Sarah E Smith, Rong Li, Kathleen Gould

    The contractile ring is a complex molecular apparatus important for dividing many eukaryotic cells. Despite knowledge of its composition, the molecular architecture of the ring is not known. Here we applied super-resolution microscopy and FRET to determine the nanoscale spatial organization of Schizosaccharomyces pombe contractile ring components relative to the plasma membrane. As in other membrane-tethered actin structures, contractile ring proteins are stratified relative to the membrane. The lowest layer (0-80 nm) contains membrane-binding scaffolds, formin, and the myosin-II tail. An intermediate zone (80-160 nm) consists of a network of cytokinesis accessory proteins and signaling components that influence cell division. Most interior from the membrane (160-400 nm) is F-actin, myosin motor domains, and an F-actin crosslinker. Circumferentially within the ring, multiple proximal membrane proteins form different sized clusters, while components farther from the membrane are uniformly distributed. This comprehensive organizational map provides a framework for understanding contractile ring function.

    更新日期:2017-09-18
  • Synaptic up-scaling preserves motor circuit output after chronic, natural inactivity
    eLife (IF 7.725) Pub Date : 2017-09-15
    Joseph M Santin, Mauricio Vallejo, Lynn K Hartzler

    Neural systems use homeostatic plasticity to maintain normal brain functions and to prevent abnormal activity. Surprisingly, homeostatic mechanisms that regulate circuit output have mainly been demonstrated during artificial and/or pathological perturbations. Natural, physiological scenarios that activate these stabilizing mechanisms in neural networks of mature animals remain elusive. To establish the extent to which a naturally inactive circuit engages mechanisms of homeostatic plasticity, we utilized the respiratory motor circuit in bullfrogs that normally remains inactive for several months during the winter. We found that inactive respiratory motoneurons exhibit a classic form of homeostatic plasticity, up-scaling of AMPA-glutamate receptors. Up-scaling increased the synaptic strength of respiratory motoneurons and acted to boost motor amplitude from the respiratory network following months of inactivity. Our results show that synaptic scaling sustains strength of the respiratory motor output following months of inactivity, thereby supporting a major neuroscience hypothesis in a normal context for an adult animal.

    更新日期:2017-09-18
  • Scc2/Nipbl hops between chromosomal cohesin rings after loading
    eLife (IF 7.725) Pub Date : 2017-09-15
    James DP Rhodes, Davide Mazza, Kim A Nasmyth, Stephan Uphoff

    The cohesin complex mediates DNA-DNA interactions both between (sister chromatid cohesion) and within chromosomes (DNA looping). It has been suggested that intra-chromosome loops are generated by extrusion of DNAs through the lumen of cohesin's ring. Scc2 (Nipbl) stimulates cohesin's ABC-like ATPase and is essential for loading cohesin onto chromosomes. However, it is possible that the stimulation of cohesin's ATPase by Scc2 also has a post-loading function, for example driving loop extrusion. Using fluorescence recovery after photobleaching (FRAP) and single- molecule tracking, we show that Scc2 binds dynamically to chromatin, principally through an association with cohesin. Scc2's movement within chromatin is consistent with a 'stop-and-go' or 'hopping' motion. We suggest that a low diffusion coefficient, a low stoichiometry relative to cohesin, and a high affinity for chromosomal cohesin enables Scc2 to move rapidly from one chromosomal cohesin complex to another, performing a function distinct from loading.

    更新日期:2017-09-18
  • Cortex-wide BOLD fMRI activity reflects locally-recorded slow oscillation-associated calcium waves
    eLife (IF 7.725) Pub Date : 2017-09-15
    Miriam Schwalm, Florian Schmid, Lydia Wachsmuth, Hendrik Backhaus, Andrea Kronfeld, Felipe Aedo Jury, Pierre-Hugues Prouvot, Consuelo Fois, Franziska Albers, Timo van Alst, Cornelius Faber, Albrecht Stroh

    Spontaneous slow oscillation-associated slow wave activity represents an internally generated state which is characterized by alternations of network quiescence and stereotypical episodes of neuronal activity - slow wave events. However, it remains unclear which macroscopic signal is related to these active periods of the slow wave rhythm. We used optic fiber-based calcium recordings of local neural populations in cortex and thalamus to detect neurophysiologically defined slow calcium waves in isoflurane anesthetized rats. The individual slow wave events were used for an event-related analysis of simultaneously acquired whole-brain BOLD fMRI. We identified BOLD responses directly related to onsets of slow calcium waves, revealing a cortex-wide BOLD correlate: the entire cortex was engaged in this specific type of slow wave activity. These findings demonstrate a direct relation of defined neurophysiological events to a specific BOLD activity pattern and were confirmed for ongoing slow wave activity by independent component and seed-based analyses.

    更新日期:2017-09-18
  • A causal role for right frontopolar cortex in directed, but not random, exploration
    eLife (IF 7.725) Pub Date : 2017-09-15
    Wojciech K Zajkowski, Malgorzata Kossut, Robert C Wilson

    The explore-exploit dilemma occurs anytime we must choose between exploring unknown options for information and exploiting known resources for reward. Previous work suggests that people use two different strategies to solve the explore-exploit dilemma: directed exploration, driven by information seeking, and random exploration, driven by decision noise. Here, we show that these two strategies rely on different neural systems. Using transcranial magnetic stimulation to inhibit the right frontopolar cortex, we were able to selectively inhibit directed exploration while leaving random exploration intact. This suggests a causal role for right frontopolar cortex in directed, but not random, exploration and that directed and random exploration rely on (at least partially) dissociable neural systems.

    更新日期:2017-09-18
  • Rift Valley fever phlebovirus NSs protein core domain structure suggests molecular basis for nuclear filaments
    eLife (IF 7.725) Pub Date : 2017-09-15
    Michal Barski, Benjamin Brennan, Ona K Miller, Jane A Potter, Swetha Vijayakrishnan, David Bhella, James H Naismith, Richard M Elliott, Ulrich Schwarz-Linek

    Rift Valley fever phlebovirus (RVFV) is a clinically and economically important pathogen increasingly likely to cause widespread epidemics. RVFV virulence depends on the interferon antagonist non-structural protein (NSs), which remains poorly characterized. We identified a stable core domain of RVFV NSs (residues 83–248), and solved its crystal structure, a novel all-helical fold organized into highly ordered fibrils. A hallmark of RVFV pathology is NSs filament formation in infected cell nuclei. Recombinant virus encoding the NSs core domain induced intranuclear filaments, suggesting it contains all essential determinants for nuclear translocation and filament formation. Mutations of key crystal fibril interface residues in viruses encoding full-length NSs completely abrogated intranuclear filament formation in infected cells. We propose the fibrillar arrangement of the NSs core domain in crystals reveals the molecular basis of assembly of this key virulence factor in cell nuclei. Our findings have important implications for fundamental understanding of RVFV virulence. https://doi.org/10.7554/eLife.29236.001

    更新日期:2017-09-18
  • Lipids and ions traverse the membrane by the same physical pathway in the nhTMEM16 scramblase
    eLife (IF 7.725) Pub Date : 2017-09-16
    Tao Jiang, Kuai Yu, H Criss Hartzell, Emad Tajkhorshid

    From bacteria to mammals, different phospholipid species are segregated between the inner and outer leaflets of the plasma membrane by ATP-dependent lipid transporters. Disruption of this asymmetry by ATP-independent phospholipid scrambling is important in cellular signaling, but its mechanism remains incompletely understood. Using MD simulations coupled with experimental assays, we show that the surface hydrophilic transmembrane cavity exposed to the lipid bilayer on the fungal scramblase nhTMEM16 serves as the pathway for both lipid translocation and ion conduction across the membrane. Ca2+ binding stimulates its open conformation by altering the structure of transmembrane helices that line the cavity. We have identified key amino acids necessary for phospholipid scrambling and validated the idea that ions permeate TMEM16 Cl- channels via a structurally homologous pathway by showing that mutation of two residues in the pore region of the TMEM16A Ca2+-activated Cl- channel convert it into a robust scramblase.

    更新日期:2017-09-18
  • Direct modulation of aberrant brain network connectivity through real-time neurofeedback
    eLife (IF 7.725) Pub Date : 2017-09-16
    Michal Ramot, Sara Kimmich, Javier Gonzalez-Castillo, Vinai Roopchansingh, Haroon Popal, Emily White, Stephen J Gotts, Alex Martin

    The existence of abnormal connectivity patterns between resting state networks in neuropsychiatric disorders, including Autism Spectrum Disorder (ASD), has been well established. Traditional treatment methods in ASD are limited, and do not address the aberrant network structure. Using real-time fMRI neurofeedback, we directly trained 3 brain nodes in participants with ASD, in which the aberrant connectivity has been shown to correlate with symptom severity. Desired network connectivity patterns were reinforced in real-time, without participants' awareness of the training taking place. This training regimen produced large, significant long-term changes in correlations at the network level, and whole brain analysis revealed that the greatest changes were focused on the areas being trained. These changes were not found in the control group. Moreover, changes in ASD resting state connectivity following the training were correlated to changes in behavior, suggesting that neurofeedback can be used to directly alter complex, clinically relevant network connectivity patterns.

    更新日期:2017-09-18
  • Lineage tracing of genome-edited alleles reveals high fidelity axolotl limb regeneration
    eLife (IF 7.725) Pub Date : 2017-09-16
    Grant Parker Flowers, Lucas D Sanor, Craig M Crews

    Salamanders are unparalleled among tetrapods in their ability to regenerate many structures, including entire limbs, and the study of this ability may provide insights into human regenerative therapies. The complex structure of the limb poses challenges to the investigation of the cellular and molecular basis of its regeneration. Using CRISPR/Cas, we genetically labelled unique cell lineages within the developing axolotl embryo and tracked the frequency of each lineage within amputated and fully regenerated limbs. This allowed us, for the first time, to assess the contributions of multiple low frequency cell lineages to the regenerating limb at once. Our comparisons reveal that regenerated limbs are high fidelity replicas of the originals even after repeated amputations.

    更新日期:2017-09-18
  • Signaling: Enzymatic insights into an inherited genetic disorder
    eLife (IF 7.725) Pub Date : 2017-09-14
    Liping Zhang, Kelly G Ten Hagen

    Mutations in an enzyme involved in protein degradation affect a signaling pathway that stimulates the development of the digestive tract.

    更新日期:2017-09-15
  • Work minimization accounts for footfall phasing in slow quadrupedal gaits
    eLife (IF 7.725) Pub Date : 2017-09-14
    James R Usherwood, Zoe T Self Davies

    Quadrupeds, like most bipeds, tend to walk with an even left/right footfall timing. However, the phasing between hind and forelimbs shows considerable variation. Here, we account for this variation by modeling and explaining the influence of hind-fore limb phasing on mechanical work requirements. These mechanics account for the different strategies used by: (1) slow animals (a group including crocodile, tortoise, hippopotamus and some babies); (2) normal medium to large mammals; and (3) (with an appropriate minus sign) sloths undertaking suspended locomotion across a range of speeds. While the unusual hind-fore phasing of primates does not match global work minimizing predictions, it does approach an only slightly more costly local minimum. Phases predicted to be particularly costly have not been reported in nature. https://doi.org/10.7554/eLife.29495.001

    更新日期:2017-09-15
  • Centriole triplet microtubules are required for stable centriole formation and inheritance in human cells
    eLife (IF 7.725) Pub Date : 2017-09-14
    Jennifer T Wang, Dong Kong, Christian R Hoerner, Jadranka Loncarek, Tim Stearns

    Centrioles are composed of long-lived microtubules arranged in nine triplets. However, the contribution of triplet microtubules to mammalian centriole formation and stability is unknown. Little is known of the mechanism of triplet microtubule formation, but experiments in unicellular eukaryotes indicate that delta-tubulin and epsilon-tubulin, two less-studied tubulin family members, are required. Here, we report that centrioles in delta-tubulin and epsilon-tubulin null mutant human cells lack triplet microtubules and fail to undergo centriole maturation. These aberrant centrioles are formed de novo each cell cycle, but are unstable and do not persist to the next cell cycle, leading to a futile cycle of centriole formation and­­­ disintegration. Disintegration can be suppressed by paclitaxel treatment. Delta-tubulin and epsilon-tubulin physically interact, indicating that these tubulins act together to maintain triplet microtubules and that these are necessary for inheritance of centrioles from one cell cycle to the next

    更新日期:2017-09-14
  • Kinetochore inactivation by expression of a repressive mRNA
    eLife (IF 7.725) Pub Date : 2017-09-14
    Jingxun Chen, Amy Tresenrider, Minghao Chia, David T McSwiggen, Gianpiero Spedale, Victoria Jorgensen, Hanna Liao, Folkert Jacobus van Werven, Elcin Unal

    Differentiation programs such as meiosis depend on extensive gene regulation to mediate cellular morphogenesis. Meiosis requires transient removal of the outer kinetochore, the complex that connects microtubules to chromosomes. How the meiotic gene expression program temporally restricts kinetochore function is unknown. We discovered that in budding yeast, kinetochore inactivation occurs by reducing the abundance of a limiting subunit, Ndc80. Furthermore, we uncovered an integrated mechanism that acts at the transcriptional and translational level to repress NDC80 expression. Central to this mechanism is the developmentally controlled transcription of an alternate NDC80 mRNA isoform, which itself cannot produce protein due to regulatory upstream ORFs in its extended 5' leader. Instead, transcription of this isoform represses the canonical NDC80 mRNA expression in cis, thereby inhibiting Ndc80 protein synthesis. This model of gene regulation raises the intriguing notion that transcription of an mRNA, despite carrying a canonical coding sequence, can directly cause gene repression.

    更新日期:2017-09-14
  • Transcription of a 5' extended mRNA isoform directs dynamic chromatin changes and interference of a downstream promoter
    eLife (IF 7.725) Pub Date : 2017-09-14
    Minghao Chia, Amy Tresenrider, Jingxun Chen, Gianpiero Spedale, Victoria Jorgensen, Elçin Ünal, Folkert Jacobus van Werven

    Cell differentiation programs require dynamic regulation of gene expression. During meiotic prophase in Saccharomyces cerevisiae, expression of the kinetochore complex subunit Ndc80 is downregulated by a 5' extended long undecoded NDC80 transcript isoform. Here we demonstrate a transcriptional interference mechanism that is responsible for inhibiting expression of the coding NDC80 mRNA isoform. Transcription from a distal NDC80 promoter directs Set1-dependent histone H3K4 dimethylation and Set2-dependent H3K36 trimethylation to establish a repressive chromatin state in the downstream canonical NDC80 promoter. As a consequence, NDC80 expression is repressed during meiotic prophase. The transcriptional mechanism described here is rapidly reversible, adaptable to fine-tune gene expression, and relies on Set2 and the Set3 histone deacetylase complex. Thus, expression of a 5' extended mRNA isoform causes transcriptional interference at the downstream promoter. We demonstrate that this is an effective mechanism to promote dynamic changes in gene expression during cell differentiation.

    更新日期:2017-09-14
  • The AAA protein Msp1 mediates clearance of excess tail-anchored proteins from the peroxisomal membrane
    eLife (IF 7.725) Pub Date : 2017-09-14
    Nicholas R Weir, Roarke A Kamber, James S Martenson, Vladimir Denic

    Msp1 is a conserved AAA ATPase in budding yeast localized to mitochondria where it prevents accumulation of mistargeted tail-anchored (TA) proteins, including the peroxisomal TA protein Pex15. Msp1 also resides on peroxisomes but it remains unknown how native TA proteins on mitochondria and peroxisomes evade Msp1 surveillance. We used live-cell quantitative cell microscopy tools and drug-inducible gene expression to dissect Msp1 function. We found that a small fraction of peroxisomal Pex15, exaggerated by overexpression, is turned over by Msp1. Kinetic measurements guided by theoretical modeling revealed that Pex15 molecules at mitochondria display age-independent Msp1 sensitivity. By contrast, Pex15 molecules at peroxisomes are rapidly converted from an initial Msp1-sensitive to an Msp1-resistant state. Lastly, we show that Pex15 interacts with the peroxisomal membrane protein Pex3, which shields Pex15 from Msp1-dependent turnover. In sum, our work argues that Msp1 selects its substrates on the basis of their solitary membrane existence.

    更新日期:2017-09-14
  • Kinetochore inactivation by expression of a repressive mRNA
    eLife (IF 7.725) Pub Date : 2017-09-14
    Jingxun Chen, Amy Tresenrider, Minghao Chia, David T McSwiggen, Gianpiero Spedale, Victoria Jorgensen, Hanna Liao, Folkert Jacobus van Werven, Elcin Unal

    Differentiation programs such as meiosis depend on extensive gene regulation to mediate cellular morphogenesis. Meiosis requires transient removal of the outer kinetochore, the complex that connects microtubules to chromosomes. How the meiotic gene expression program temporally restricts kinetochore function is unknown. We discovered that in budding yeast, kinetochore inactivation occurs by reducing the abundance of a limiting subunit, Ndc80. Furthermore, we uncovered an integrated mechanism that acts at the transcriptional and translational level to repress NDC80 expression. Central to this mechanism is the developmentally controlled transcription of an alternate NDC80 mRNA isoform, which itself cannot produce protein due to regulatory upstream ORFs in its extended 5' leader. Instead, transcription of this isoform represses the canonical NDC80 mRNA expression in cis, thereby inhibiting Ndc80 protein synthesis. This model of gene regulation raises the intriguing notion that transcription of an mRNA, despite carrying a canonical coding sequence, can directly cause gene repression.

    更新日期:2017-09-14
  • Centriole triplet microtubules are required for stable centriole formation and inheritance in human cells
    eLife (IF 7.725) Pub Date : 2017-09-14
    Jennifer T Wang, Dong Kong, Christian R Hoerner, Jadranka Loncarek, Tim Stearns

    Centrioles are composed of long-lived microtubules arranged in nine triplets. However, the contribution of triplet microtubules to mammalian centriole formation and stability is unknown. Little is known of the mechanism of triplet microtubule formation, but experiments in unicellular eukaryotes indicate that delta-tubulin and epsilon-tubulin, two less-studied tubulin family members, are required. Here, we report that centrioles in delta-tubulin and epsilon-tubulin null mutant human cells lack triplet microtubules and fail to undergo centriole maturation. These aberrant centrioles are formed de novo each cell cycle, but are unstable and do not persist to the next cell cycle, leading to a futile cycle of centriole formation and­­­ disintegration. Disintegration can be suppressed by paclitaxel treatment. Delta-tubulin and epsilon-tubulin physically interact, indicating that these tubulins act together to maintain triplet microtubules and that these are necessary for inheritance of centrioles from one cell cycle to the next

    更新日期:2017-09-14
  • The AAA protein Msp1 mediates clearance of excess tail-anchored proteins from the peroxisomal membrane
    eLife (IF 7.725) Pub Date : 2017-09-14
    Nicholas R Weir, Roarke A Kamber, James S Martenson, Vladimir Denic

    Msp1 is a conserved AAA ATPase in budding yeast localized to mitochondria where it prevents accumulation of mistargeted tail-anchored (TA) proteins, including the peroxisomal TA protein Pex15. Msp1 also resides on peroxisomes but it remains unknown how native TA proteins on mitochondria and peroxisomes evade Msp1 surveillance. We used live-cell quantitative cell microscopy tools and drug-inducible gene expression to dissect Msp1 function. We found that a small fraction of peroxisomal Pex15, exaggerated by overexpression, is turned over by Msp1. Kinetic measurements guided by theoretical modeling revealed that Pex15 molecules at mitochondria display age-independent Msp1 sensitivity. By contrast, Pex15 molecules at peroxisomes are rapidly converted from an initial Msp1-sensitive to an Msp1-resistant state. Lastly, we show that Pex15 interacts with the peroxisomal membrane protein Pex3, which shields Pex15 from Msp1-dependent turnover. In sum, our work argues that Msp1 selects its substrates on the basis of their solitary membrane existence.

    更新日期:2017-09-14
  • Transcription of a 5' extended mRNA isoform directs dynamic chromatin changes and interference of a downstream promoter
    eLife (IF 7.725) Pub Date : 2017-09-14
    Minghao Chia, Amy Tresenrider, Jingxun Chen, Gianpiero Spedale, Victoria Jorgensen, Elçin Ünal, Folkert Jacobus van Werven

    Cell differentiation programs require dynamic regulation of gene expression. During meiotic prophase in Saccharomyces cerevisiae, expression of the kinetochore complex subunit Ndc80 is downregulated by a 5' extended long undecoded NDC80 transcript isoform. Here we demonstrate a transcriptional interference mechanism that is responsible for inhibiting expression of the coding NDC80 mRNA isoform. Transcription from a distal NDC80 promoter directs Set1-dependent histone H3K4 dimethylation and Set2-dependent H3K36 trimethylation to establish a repressive chromatin state in the downstream canonical NDC80 promoter. As a consequence, NDC80 expression is repressed during meiotic prophase. The transcriptional mechanism described here is rapidly reversible, adaptable to fine-tune gene expression, and relies on Set2 and the Set3 histone deacetylase complex. Thus, expression of a 5' extended mRNA isoform causes transcriptional interference at the downstream promoter. We demonstrate that this is an effective mechanism to promote dynamic changes in gene expression during cell differentiation.

    更新日期:2017-09-14
  • Distinct stages of synapse elimination are induced by burst firing of CA1 neurons and differentially require MEF2A/D
    eLife (IF 7.725) Pub Date : 2017-09-13
    Chia-Wei Chang, Julia Wilkerson, Carly Hale, Jay R Gibson, Kimberly M Huber

    Experience and activity refine cortical circuits through synapse elimination, but little is known about the activity patterns and downstream molecular mechanisms that mediate this process. We used optogenetics to drive individual mouse CA1 hippocampal neurons to fire in theta frequency bursts to understand how cell autonomous, postsynaptic activity leads to synapse elimination. Brief (1hr) periods of postsynaptic bursting selectively depressed AMPA receptor (R) synaptic transmission, or silenced excitatory synapses, whereas more prolonged (24 hr) firing depressed both AMPAR and NMDAR EPSCs and eliminated spines, indicative of a synapse elimination. Both synapse silencing and elimination required de novo transcription, but only silencing required the activity-dependent transcription factors MEF2A/D. Burst firing induced MEF2A/D-dependent induction of the target gene Arc which contributed to synapse silencing and elimination. This work reveals new and distinct forms of activity-dependent synapse depression and suggests that these processes can occur independently.

    更新日期:2017-09-13
  • Biochemical adaptations of the retina and retinal pigment epithelium support a metabolic ecosystem in the vertebrate eye
    eLife (IF 7.725) Pub Date : 2017-09-13
    Mark A Kanow, Michelle M Giarmarco, Connor SR Jankowski, Kristine Tsantilas, Abbi L Engel, Jianhai Du, Jonathan D Linton, Christopher C Farnsworth, Stephanie R Sloat, Austin Rountree, Ian R Sweet, Ken J Lindsay, Edward D Parker, Susan E Brockerhoff, Martin Sadilek, Jennifer R Chao, James B Hurley

    Here we report multiple lines of evidence for a comprehensive model of energy metabolism in the vertebrate eye. Metabolic flux, locations of key enzymes, and our finding that glucose enters mouse and zebrafish retinas mostly through photoreceptors support a conceptually new model for retinal metabolism. In this model, glucose from the choroidal blood passes through the retinal pigment epithelium to the retina where photoreceptors convert it to lactate. Photoreceptors then export the lactate as fuel for the retinal pigment epithelium and for neighboring Müller glial cells. We used human retinal epithelial cells to show that lactate can suppress consumption of glucose by the retinal pigment epithelium. Suppression of glucose consumption in the retinal pigment epithelium can increase the amount of glucose that reaches the retina. This framework for understanding metabolic relationships in the vertebrate retina provides new insights into the underlying causes of retinal disease and age-related vision loss.

    更新日期:2017-09-13
  • A case study for a psychographic-behavioral segmentation approach for targeted demand generation in voluntary medical male circumcision
    eLife (IF 7.725) Pub Date : 2017-09-13
    Sema K Sgaier, Maria Eletskaya, Elisabeth Engl, Owen Mugurungi, Bushimbwa Tambatamba, Gertrude Ncube, Sinokuthemba Xaba, Alice Nanga, Svetlana Gogolina, Patrick Odawo, Sehlulekile Gumede-Moyo, Steve Kretschmer

    Public health programs are starting to recognize the need to move beyond a one-size-fits-all approach in demand generation, and instead tailor interventions to the heterogeneity underlying human decision making. Currently, however, there is a lack of methods to enable such targeting. We describe a novel hybrid behavioral-psychographic segmentation approach to segment stakeholders on potential barriers to a target behavior. We then apply the method in a case study of demand generation for voluntary medical male circumcision (VMMC) among 15-29-year-old males in Zambia and Zimbabwe. Canonical correlations and hierarchical clustering techniques were applied on representative samples of men in each country who were differentiated by their underlying reasons for their propensity to get circumcised. We characterized six distinct segments of men in Zimbabwe, and seven segments in Zambia, according to their needs, perceptions, attitudes and behaviors towards VMMC, thus highlighting distinct reasons for a failure to engage in the desired behavior.

    更新日期:2017-09-13
  • Stone Tool Use: Monkeys overharvest shellfish
    eLife (IF 7.725) Pub Date : 2017-09-13
    George H Perry, Brian F Codding

    The use of stone tools by macaques in Thailand has reduced the size and population density of coastal shellfish: previously it was thought that overharvesting effects resulted from human activity alone.

    更新日期:2017-09-13
  • Causal evidence for lateral prefrontal cortex dynamics supporting cognitive control
    eLife (IF 7.725) Pub Date : 2017-09-13
    Derek Evan Nee, Mark D'Esposito

    The lateral prefrontal cortex (LPFC) is essential for higher-level cognition, but how its interactions support cognitive control remains elusive. Previously (Nee and D'Esposito, 2016), dynamic causal modeling (DCM) indicated that mid LPFC integrates abstract, rostral and concrete, caudal influences to inform context-appropriate action. Here, we use continuous theta-burst transcranial magnetic stimulation (cTBS) to causally test this model. cTBS was applied to three LPFC sites and a control site in counterbalanced sessions. Behavioral modulations resulting from cTBS were largely predicted by information flow within the previously estimated DCM. However, cTBS to caudal LPFC unexpectedly impaired processes presumed to involve rostral LPFC. Adding a pathway from caudal to mid-rostral LPFC significantly improved the model fit and accounted for the observed behavioral findings. These data provide causal evidence for LPFC dynamics supporting cognitive control and demonstrate the utility of combining DCM with causal manipulations to test and refine models of cognition.

    更新日期:2017-09-13
  • Multiple conserved cell adhesion protein interactions mediate neural wiring of a sensory circuit in C. elegans
    eLife (IF 7.725) Pub Date : 2017-09-13
    Byunghyuk Kim, Scott W Emmons

    Nervous system function relies on precise synaptic connections. A number of widely-conserved cell adhesion proteins are implicated in cell recognition between synaptic partners, but how these proteins act as a group to specify a complex neural network is poorly understood. Taking advantage of known connectivity in C. elegans, we identified and studied cell adhesion genes expressed in three interacting neurons in the mating circuits of the adult male. Two interacting pairs of cell surface proteins independently promote fasciculation between sensory neuron HOA and its postsynaptic target interneuron AVG: BAM-2/neurexin-related in HOA binds to CASY-1/calsyntenin in AVG; SAX-7/L1CAM in sensory neuron PHC binds to RIG-6/contactin in AVG. A third, basal pathway results in considerable HOA-AVG fasciculation and synapse formation in the absence of the other two. The features of this multiplexed mechanism help to explain how complex connectivity is encoded and robustly established during nervous system development.

    更新日期:2017-09-13
  • Causal evidence for lateral prefrontal cortex dynamics supporting cognitive control
    eLife (IF 7.725) Pub Date : 2017-09-13
    Derek Evan Nee, Mark D'Esposito

    The lateral prefrontal cortex (LPFC) is essential for higher-level cognition, but how its interactions support cognitive control remains elusive. Previously (Nee and D'Esposito, 2016), dynamic causal modeling (DCM) indicated that mid LPFC integrates abstract, rostral and concrete, caudal influences to inform context-appropriate action. Here, we use continuous theta-burst transcranial magnetic stimulation (cTBS) to causally test this model. cTBS was applied to three LPFC sites and a control site in counterbalanced sessions. Behavioral modulations resulting from cTBS were largely predicted by information flow within the previously estimated DCM. However, cTBS to caudal LPFC unexpectedly impaired processes presumed to involve rostral LPFC. Adding a pathway from caudal to mid-rostral LPFC significantly improved the model fit and accounted for the observed behavioral findings. These data provide causal evidence for LPFC dynamics supporting cognitive control and demonstrate the utility of combining DCM with causal manipulations to test and refine models of cognition.

    更新日期:2017-09-13
  • A case study for a psychographic-behavioral segmentation approach for targeted demand generation in voluntary medical male circumcision
    eLife (IF 7.725) Pub Date : 2017-09-13
    Sema K Sgaier, Maria Eletskaya, Elisabeth Engl, Owen Mugurungi, Bushimbwa Tambatamba, Gertrude Ncube, Sinokuthemba Xaba, Alice Nanga, Svetlana Gogolina, Patrick Odawo, Sehlulekile Gumede-Moyo, Steve Kretschmer

    Public health programs are starting to recognize the need to move beyond a one-size-fits-all approach in demand generation, and instead tailor interventions to the heterogeneity underlying human decision making. Currently, however, there is a lack of methods to enable such targeting. We describe a novel hybrid behavioral-psychographic segmentation approach to segment stakeholders on potential barriers to a target behavior. We then apply the method in a case study of demand generation for voluntary medical male circumcision (VMMC) among 15-29-year-old males in Zambia and Zimbabwe. Canonical correlations and hierarchical clustering techniques were applied on representative samples of men in each country who were differentiated by their underlying reasons for their propensity to get circumcised. We characterized six distinct segments of men in Zimbabwe, and seven segments in Zambia, according to their needs, perceptions, attitudes and behaviors towards VMMC, thus highlighting distinct reasons for a failure to engage in the desired behavior.

    更新日期:2017-09-13
  • Biochemical adaptations of the retina and retinal pigment epithelium support a metabolic ecosystem in the vertebrate eye
    eLife (IF 7.725) Pub Date : 2017-09-13
    Mark A Kanow, Michelle M Giarmarco, Connor SR Jankowski, Kristine Tsantilas, Abbi L Engel, Jianhai Du, Jonathan D Linton, Christopher C Farnsworth, Stephanie R Sloat, Austin Rountree, Ian R Sweet, Ken J Lindsay, Edward D Parker, Susan E Brockerhoff, Martin Sadilek, Jennifer R Chao, James B Hurley

    Here we report multiple lines of evidence for a comprehensive model of energy metabolism in the vertebrate eye. Metabolic flux, locations of key enzymes, and our finding that glucose enters mouse and zebrafish retinas mostly through photoreceptors support a conceptually new model for retinal metabolism. In this model, glucose from the choroidal blood passes through the retinal pigment epithelium to the retina where photoreceptors convert it to lactate. Photoreceptors then export the lactate as fuel for the retinal pigment epithelium and for neighboring Müller glial cells. We used human retinal epithelial cells to show that lactate can suppress consumption of glucose by the retinal pigment epithelium. Suppression of glucose consumption in the retinal pigment epithelium can increase the amount of glucose that reaches the retina. This framework for understanding metabolic relationships in the vertebrate retina provides new insights into the underlying causes of retinal disease and age-related vision loss.

    更新日期:2017-09-13
  • Distinct stages of synapse elimination are induced by burst firing of CA1 neurons and differentially require MEF2A/D
    eLife (IF 7.725) Pub Date : 2017-09-13
    Chia-Wei Chang, Julia Wilkerson, Carly Hale, Jay R Gibson, Kimberly M Huber

    Experience and activity refine cortical circuits through synapse elimination, but little is known about the activity patterns and downstream molecular mechanisms that mediate this process. We used optogenetics to drive individual mouse CA1 hippocampal neurons to fire in theta frequency bursts to understand how cell autonomous, postsynaptic activity leads to synapse elimination. Brief (1hr) periods of postsynaptic bursting selectively depressed AMPA receptor (R) synaptic transmission, or silenced excitatory synapses, whereas more prolonged (24 hr) firing depressed both AMPAR and NMDAR EPSCs and eliminated spines, indicative of a synapse elimination. Both synapse silencing and elimination required de novo transcription, but only silencing required the activity-dependent transcription factors MEF2A/D. Burst firing induced MEF2A/D-dependent induction of the target gene Arc which contributed to synapse silencing and elimination. This work reveals new and distinct forms of activity-dependent synapse depression and suggests that these processes can occur independently.

    更新日期:2017-09-13
  • Multiple conserved cell adhesion protein interactions mediate neural wiring of a sensory circuit in C. elegans
    eLife (IF 7.725) Pub Date : 2017-09-13
    Byunghyuk Kim, Scott W Emmons

    Nervous system function relies on precise synaptic connections. A number of widely-conserved cell adhesion proteins are implicated in cell recognition between synaptic partners, but how these proteins act as a group to specify a complex neural network is poorly understood. Taking advantage of known connectivity in C. elegans, we identified and studied cell adhesion genes expressed in three interacting neurons in the mating circuits of the adult male. Two interacting pairs of cell surface proteins independently promote fasciculation between sensory neuron HOA and its postsynaptic target interneuron AVG: BAM-2/neurexin-related in HOA binds to CASY-1/calsyntenin in AVG; SAX-7/L1CAM in sensory neuron PHC binds to RIG-6/contactin in AVG. A third, basal pathway results in considerable HOA-AVG fasciculation and synapse formation in the absence of the other two. The features of this multiplexed mechanism help to explain how complex connectivity is encoded and robustly established during nervous system development.

    更新日期:2017-09-13
Some contents have been Reproduced with permission of the American Chemical Society.
Some contents have been Reproduced by permission of The Royal Society of Chemistry.
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