Skip to main content
SpringerLink
Log in

Genome-wide identification and expression profiles of phased siRNAs in a male-sterile somatic cybrid of pummelo (Citrus grandis)

  • Original Article
  • Published:
Tree Genetics & Genomes Aims and scope Submit manuscript

Abstract

Phased small interfering RNAs (phasiRNAs) are 21- or 24-nucleotide small non-coding RNAs that play important regulatory roles in various biological processes of plant growth and development. Biogenesis of phasiRNAs is typically triggered by miRNAs and involves dicer-processing. In our previous study, we constructed high-throughput sRNA and degradome sequencing libraries out of developing flower of the male sterile cybrid pummelo (G1 + HBP) and its fertile protoplast fusion parent Hirado Buntan pummelo (HBP), and identified the miRNAs involved in citrus male sterility. In this study, 280 PHAS loci generating 2235 unique 21-nucleotide phasiRNAs were identified in the flowers of the cybrid pummelo and its fertile fusion parent; of them, 210 overlapped with annotated protein coding genes, and 26 encoded transcription factors families involved in plant development, including ARF, MYB, AP2, NAC, GRF, TIR1/AFB, and AGL. Based on integration of miRNAs, phasiRNA and the degradome data, we identified nine miRNAs and eight phasiRNAs that trigger phasiRNAs production out of 29 PHAS loci, and 67 genes of broad regulatory activities were identified to be targets of phasiRNAs. We further identified nine miRNA-PHAS-phasiRNA-targets regulatory cascades, one phasiRNA-PHAS-phasiRNA-targets cascade, and 88 phasiRNAs with differential expression patterns in flowers between the cybrid and HBP. This analysis provided new information about miRNA-PHAS-phasiRNA-target networks and suggested potential roles of phasiRNAs in male sterility of pummelo and contributed to further study of phasiRNA function in citrus.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Addo-Quaye C, Miller W, Axtell MJ (2009) CleaveLand: a pipeline for using degradome data to find cleaved small RNA targets. Bioinformatics 25:130–131

    CAS  PubMed  Google Scholar 

  • Allen E, Howell MD (2010) miRNAs in the biogenesis of trans-acting siRNAs in higher plants. Semin Cell Dev Biol 21:798–804

    CAS  PubMed  Google Scholar 

  • Allen E, Xie Z, Gustafson AM, Carrington JC (2005) microRNA-directed phasing during trans-acting siRNA biogenesis in plants. Cell 121:207–221

    CAS  PubMed  Google Scholar 

  • Arikit S, Xia R, Kakrana A, Huang K, Zhai J, Yan Z, Valdés-López O, Prince S, Musket TA, Nguyen HT (2014) An atlas of soybean small RNAs identifies phased siRNAs from hundreds of coding genes. Plant Cell 26:4584–4601

    CAS  PubMed  PubMed Central  Google Scholar 

  • Axtell MJ, Jan C, Rajagopalan R, Bartel DP (2006) A two-hit trigger for siRNA biogenesis in plants. Cell 127:565–577

    CAS  PubMed  Google Scholar 

  • Axtell MJ (2010) A method to discover phased siRNA loci. Methods Mol Biol 592:59–70

    CAS  PubMed  Google Scholar 

  • Barkan A, Small I (2014) Pentatricopeptide repeat proteins in plants. Annu Rev Plant Biol 65:415–442

    CAS  PubMed  Google Scholar 

  • Berninger P, Gaidatzis D, Nimwegen E, Zavolan M (2008) Computational analysis of small RNA cloning data. Methods 44:13–21

    CAS  PubMed  Google Scholar 

  • Chen HM, Chen LT, Patel K, Li YH, Baulcombe DC, Wu SH (2010) 22-Nucleotide RNAs trigger secondary siRNA biogenesis in plants. Proc Natl Acad Sci 107:15269–15274

    CAS  PubMed  PubMed Central  Google Scholar 

  • Dukowic-Schulze S, Sundararajan A, Ramaraj T, Kianian S, Pawlowski WP, Mudge J, Chen C (2016) Novel meiotic miRNAs and indications for a role of phasiRNAs in meiosis. Front Plant Sci 7:762

    PubMed  PubMed Central  Google Scholar 

  • Fahlgren N, Montgomery TA, Howell MD, Allen E, Dvorak SK, Alexander AL, Carrington JC (2006) Regulation of AUXIN RESPONSE FACTOR3 by TAS3 ta-siRNA affects developmental timing and patterning in Arabidopsis. Curr Biol 16:939–944

    CAS  PubMed  Google Scholar 

  • Fan Y, Yang J, Mathioni SM, Yu J, Shen J, Yang X, Wang L, Zhang Q, Cai Z, Xu C, Li X, Xiao J, Meyers BC, Zhang Q (2016) PMS1T, producing phased small-interfering RNAs, regulates photoperiod-sensitive male sterility in rice. Proc Natl Acad Sci 113:15144–15149

    CAS  PubMed  PubMed Central  Google Scholar 

  • Fang YN, Qiu WM, Wang Y, Wu XM, Xu Q, Guo WW (2014) Identification of differentially expressed microRNAs from a male sterile Ponkan mandarin (Citrus reticulata Blanco) and its fertile wild type by small RNA and degradome sequencing. Tree Genet Genoms 10:1567–1581

    Google Scholar 

  • Fang YN, Zheng BB, Wang L, Yang W, Wu XM, Xu Q, Guo WW (2016) High-throughput sequencing and degradome analysis reveal altered expression of miRNAs and their targets in a male-sterile cybrid pummelo (Citrus grandis). BMC Genomics 17:591

    PubMed  PubMed Central  Google Scholar 

  • Fei Q, Li P, Teng C, Meyers BC (2015) Secondary siRNAs from medicago NB-LRRs modulated via miRNA-target interactions and their abundances. Plant J 83:451–465

    CAS  PubMed  Google Scholar 

  • Fei Q, Xia R, Meyers BC (2013) Phased, secondary, small interfering RNAs in posttranscriptional regulatory networks. Plant Cell 25:2400–2415

    CAS  PubMed  PubMed Central  Google Scholar 

  • Garg V, Khan AW, Kudapa H, Kale SM, Chitikineni A, Qiwei S, Kishor PK (2019) Integrated transcriptome, small RNA and degradome sequencing approaches provide insights into Ascochyta blight resistance in chickpea. Plant Biotechnol J 17:914–931

    CAS  PubMed  Google Scholar 

  • Guo QL, Qu XF, Jin WB (2014) PhaseTank: genome-wide computational identification of phasiRNAs and their regulatory cascades. Bioinformatics 31:284–286

    PubMed  Google Scholar 

  • Guo WW, Prasad D, Cheng YJ, Serrano P, Deng XX, Grosser JW (2004) Targeted cybridization in citrus: transfer of Satsuma cytoplasm to seedy cultivars for potential seedlessness. Plant Cell Rep 22:752–758

    CAS  PubMed  Google Scholar 

  • Howell MD, Fahlgren N, Chapman EJ, Cumbie JS, Sullivan CM, Givan SA, Kasschau KD, Carrington JC (2007) Genome-wide analysis of the RNA-DEPENDENT RNA POLYMERASE6/DICER-LIKE4 pathway in Arabidopsis reveals dependency on miRNA- and tasiRNA-directed targeting. Plant Cell 19:926–942

    CAS  PubMed  PubMed Central  Google Scholar 

  • Hu Z, Tong Z, Wei J, Yi H, Deng X (2006) Mitochondrial gene expression in stamens is differentially regulated during male gametogenesis in Citrus unshiu. J Hortic Sci Biotechnol 81:565–569

    CAS  Google Scholar 

  • Johnson C, Kasprzewska A, Tennessen K, Fernandes J, Nan GL, Walbot V, Sundaresan V, Vance V, Bowman LH (2009) Clusters and superclusters of phased small RNAs in the developing inflorescence of rice. Genome Res 19:1429–1440

    CAS  PubMed  PubMed Central  Google Scholar 

  • Kume K, Tsutsumi K, Saitoh Y (2010) TAS1 trans-acting siRNA targets are differentially regulated at low temperature, and TAS1 trans-acting siRNA mediates temperature-controlled At1g51670 expression. Biosci Biotechnol Biochem 74:1435–1440

    CAS  PubMed  Google Scholar 

  • Langmead B, Salzberg SL (2012) Fast gapped-read alignment with Bowtie 2. Nat Methods 9:357–359

    CAS  PubMed  PubMed Central  Google Scholar 

  • Liu X, Hao L, Li D, Zhu L, Hu S (2015) Long non-coding RNAs and their biological roles in plants. Biosci Biotechnol Biochem 13:137–147

    CAS  Google Scholar 

  • Liu Y, Ke L, Wu G, Xu Y, Wu X, Xia R, Deng X, Xu Q (2017) miR3954 is a trigger of phasiRNAs that affects flowering time in citrus. Plant J 92:263–275

    CAS  PubMed  Google Scholar 

  • Liu Z, Shi X, Li S, Hu G, Zhang L, Song X (2018) Tapetal-delayed programmed cell death (PCD) and oxidative stress-induced male sterility of Aegilops uniaristata cytoplasm in wheat. Int J Mol Sci 19:1708

    PubMed Central  Google Scholar 

  • Robinson MD, McCarthy DJ, Smyth GK (2010) edgeR: a bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics 26:139–140

    CAS  PubMed  Google Scholar 

  • Rovira AG, Smith AG (2019) PPR proteins - orchestrators of organelle RNA metabolism. Physiol Plantarum 166:451–459

    CAS  Google Scholar 

  • Shivaprasad PV, Chen HM, Patel K, Bond DM, Santos BA, Baulcombe DC (2012) A microRNA superfamily regulates nucleotide binding site-leucine-rich repeats and other mRNAs. Plant Cell 24:859–874

    CAS  PubMed  PubMed Central  Google Scholar 

  • Shuai P, Su Y, Liang D, Zhang Z, Xia X, Yin W (2016) Identification of phasiRNAs and their drought- responsiveness in Populus trichocarpa. FEBS Lett 590:3616–3627

    CAS  PubMed  Google Scholar 

  • Song X, Li P, Zhai J, Zhou M, Ma L, Liu B, Jeong DH, Nakano M, Cao S, Liu C, Chu C, Wang XJ, Green PJ, Meyers BC, Cao X (2012) Roles of DCL4 and DCL3b in rice phased small RNA biogenesis. Plant J 69:462–474

    CAS  PubMed  Google Scholar 

  • Su Y, Li HG, Wang Y, Li S, Wang HL, Yu L, Li S, Wang HL, Yu L, He F, Yang Y, Feng CH, Shuai P, Liu C, Yin W, Xia X (2018) Poplar miR472a targeting NBS-LRRs is involved in effective defense against the necrotrophic fungus Cytospora chrysosperma. J Exp Bot 69:5519–5530

    CAS  PubMed  Google Scholar 

  • Tian T, Liu Y, Yan H, You Q, Yi X, Du Z, Xu W, Su Z (2017) agriGO v2. 0: a GO analysis toolkit for the agricultural community, 2017 update. Nucleic Acids Res 45:W122–W129

    CAS  PubMed  PubMed Central  Google Scholar 

  • Vazquez F, Vaucheret H, Rajagopalan R, Lepers C, Gasciolli V, Mallory AC, Hilbert JL, Bartel B, Crete P (2004) Endogenous trans-acting siRNAs regulate the accumulation of Arabidopsis mRNAs. Mol Cell 16:69–79

    CAS  PubMed  Google Scholar 

  • Visser M, van der Walt AP, Maree HJ, Rees DJG, Burger JT (2014) Extending the sRNAome of apple by next-generation sequencing. PLoS One 9:e95782

    PubMed  PubMed Central  Google Scholar 

  • Wang C, Leng X, Zhang Y, Kayesh E, Zhang Y, Sun X, Fang J (2014) Transcriptome-wide analysis of dynamic variations in regulation modes of grapevine microRNAs on their target genes during grapevine development. Plant Mol Biol 84:269–285

    CAS  PubMed  Google Scholar 

  • Wang L, Pan Z, Guo W (2010) Proteomic analysis of leaves from a diploid cybrid produced by protoplast fusion between Satsuma mandarin and pummelo. Plant Cell Tiss Org 103:165–174

    CAS  Google Scholar 

  • Wang L, Zhao S, Gu C, Zhou Y, Zhou H, Ma J, Cheng J, Han Y (2013) Deep RNA-Seq uncovers the peach transcriptome landscape. Plant Mol Biol 83:365–377

    CAS  PubMed  Google Scholar 

  • Wilson ZA, Song J, Taylor B, Yang C (2011) The final split: the regulation of anther dehiscence. J Exp Bot 62:1633–1649

    CAS  PubMed  Google Scholar 

  • Wu F, Chen Y, Tian X, Zhu X, Jin W (2017) Genome-wide identification and characterization of phased small interfering RNA genes in response to Botrytis cinerea infection in Solanum lycopersicum. Sci Rep 7:3019

    PubMed  PubMed Central  Google Scholar 

  • Wu J (2018) miRNAs as a secret weapon in the battlefield of haustoria, the interface between parasites and host plants. Mol Plant 11:354–356

    CAS  PubMed  Google Scholar 

  • Xia R, Xu J, Arikit S, Meyers BC (2015) Extensive families of miRNAs and PHAS loci in Norway spruce demonstrate the origins of complex phasiRNA networks in seed plants. Molecular Biol Evol 32:2905–2918

    CAS  Google Scholar 

  • Xia R, Zhu H, An YQ, Beers EP, Liu ZR (2012) Apple miRNAs and tasiRNAs with novel regulatory networks. Genome Biol 13:R47

    CAS  PubMed  PubMed Central  Google Scholar 

  • Yamamoto M, Matsumoto R, Okudai N, Yamada Y (1997) Aborted anthers of citrus result from gene-cytoplasmic male sterility. Sci Hort 70:9–14

    Google Scholar 

  • Yoshikawa M, Peragine A, Park MY, Poethig RS (2005) A pathway for the biogenesis of trans-acting siRNAs in Arabidopsis. Genes Dev 19:2164–2175

    CAS  PubMed  PubMed Central  Google Scholar 

  • Zhai J, Jeong DH, De Paoli E, Park S, Rosen BD, Li Y, Gonzalez AJ, Yan Z, Kitto SL, Grusak MA, Jackson SA, Stacey G, Cook DR, Green PJ, Sherrier DJ, Meyers BC (2011) MicroRNAs as master regulators of the plant NB-LRR defense gene family via the production of phased, trans-acting siRNAs. Genes Dev 25:2540–2553

    CAS  PubMed  PubMed Central  Google Scholar 

  • Zhai JX, Zhang H, Arikit S, Huang K, Nan GL, Walbot V, Meyers BC (2015) Spatiotemporally dynamic, cell-type-dependent premeiotic and meiotic phasiRNAs in maize anthers. Proc Natl Acad Sci 112:3146–3151

    CAS  PubMed  PubMed Central  Google Scholar 

  • Zheng BB, Fang YN, Pan ZY, Sun L, Deng XX, Grosser JW, Guo WW (2014) iTRAQ-based quantitative proteomics analysis revealed alterations of carbohydrate metabolism pathways and mitochondrial proteins in a male sterile cybrid pummelo. J Proteome Res 13:2998–3015

    CAS  PubMed  Google Scholar 

  • Zheng Y, Wang Y, Wu J, Ding B, Fei Z (2015) A dynamic evolutionary and functional landscape of plant phased small interfering RNAs. BMC Biol 13:32

    PubMed  PubMed Central  Google Scholar 

  • Zhu H, Xia R, Zhao BY, An YQ, Dardick CD, Callahan AM, Liu ZR (2012) Unique expression, processing regulation, and regulatory network of peach (Prunus persica) miRNAs. BMC Plant Biol 12:149

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Data archiving statement

The data of 12 sRNA libraries and one degradome library can be found in the public database Gene Expression Omnibus (http://www.ncbi.nlm.nih.gov/geo/, public on Jan 09, 2017) under the accession number of GSE76668 (Fang et al. 2016).

Funding

This research was financially supported by the National Natural Science Foundation of China (nos. 31530065, 31701891) and the Fundamental Research Funds for Central Universities (nos. 2662018PY007, 2662018PY013).

Author information

Authors and Affiliations

  1. Key Laboratory of Horticultural Plant Biology (Ministry of Education), Huazhong Agricultural University, Wuhan, 430070, China

    Yan-Ni Fang, Xiao-Ming Yang, Nan Jiang, Xiao-Meng Wu & Wen-Wu Guo

  2. Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing, 210037, China

    Xiao-Ming Yang

Authors
  1. Yan-Ni Fang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xiao-Ming Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Nan Jiang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Xiao-Meng Wu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Wen-Wu Guo
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Xiao-Meng Wu.

Additional information

Communicated by A.M. Dandekar

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Supplementary figure 1

Heat map of total abundance patterns of phasiRNAs generated from 11 PHAS loci annotated as PPR genes. The bar represents normalized log2 ratio of RPM (reads per million) value for total abundance of phasiRNAs generated on each PHAS loci. The red color indicates high abundance, and blue color indicates low abundance. HBP, Hirado Buntan pummelo (C. grandis Osbeck). G1+HBP, cybrid between HBP and “Guoqing No.1” Satsuma mandarin (C. unshiu). -1, stage 1 (from floral bud differentiation to pollen mother cell formation). -2, stage 2 (from tetrads formation to microspores release). -3, stage 3 (from young microspore development to mature pollen release). (JPG 222 kb)

ESM 1

(TIF 222 kb)

ESM 2

(XLSX 390 kb)

ESM 3

(XLSX 34 kb)

ESM 4

(XLSX 36 kb)

ESM 5

(XLSX 36 kb)

ESM 6

(XLSX 11 kb)

ESM 7

(XLSX 14 kb)

ESM 8

(XLSX 16 kb)

ESM 9

(DOCX 15 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Fang, YN., Yang, XM., Jiang, N. et al. Genome-wide identification and expression profiles of phased siRNAs in a male-sterile somatic cybrid of pummelo (Citrus grandis). Tree Genetics & Genomes 16, 46 (2020). https://doi.org/10.1007/s11295-020-01437-z

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1007/s11295-020-01437-z

Keywords

Search

Navigation