Skip to main content
SpringerLink
Log in

Genome-wide analysis of alternative splicing divergences between Brassica hexaploid and its parents

  • Original Article
  • Published:
Planta Aims and scope Submit manuscript

Abstract

Main conclusion

Compared with its parents, Brassica hexaploid underwent significant AS changes, which may provide diversified gene expression regulation patterns and could enhance its adaptability during evolution

Abstract

Polyploidization is considered a significant evolution force that promotes species formation. Alternative splicing (AS) plays a crucial role in multiple biological processes during plant growth and development. To explore the effects of allopolyploidization on the AS patterns of genes, a genome-wide AS analysis was performed by RNA-seq in Brassica hexaploid and its parents. In total, we found 7913 (27540 AS events), 14447 (70179 AS events), and 13205 (60804 AS events) AS genes in Brassica rapa, Brassica carinata, and Brassica hexaploid, respectively. A total of 920 new AS genes were discovered in Brassica hexaploid. There were 56 differently spliced genes between Brassica hexaploid and its parents. In addition, most of the alternative 5ʹ splice sites were located 4 bp upstream of the dominant 5ʹ splice sites, and most of the alternative 3ʹ splice sites were located 3 bp downstream of the dominant 3ʹ splice sites in Brassica hexapliod, which was similar to B. carinata. Furthermore, we cloned and sequenced all amplicons from the RT-PCR products of GRP7/8, namely, Bol045859, Bol016025 and Bol02880. The three genes were found to produce AS transcripts in a new way. The AS patterns of genes were diverse between Brassica hexaploid and its parents, including the loss and gain of AS events. Allopolyploidization changed alternative splicing sites of pre-mRNAs in Brassica hexaploid, which brought about alterations in the sequences of transcripts. Our study provided novel insights into the AS patterns of genes in allopolyploid plants, which may provide a reference for the study of polyploidy adaptability.

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.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9
Fig. 10
Fig. 11
Fig. 12
Fig. 13
Fig. 14

Similar content being viewed by others

Abbreviations

Alt5ʹSS:

Alternative donor sites

Alt3ʹSS:

Alternative acceptor sites

AS:

Alternative splicing

DEG:

Differentially expressed genes

DSG:

Differentially spliced genes

GO:

Gene ontology

IR:

Intron retention

NI:

New intron

PTC:

Premature termination codon

RRM:

RNA recognition domain

References

  • Acimovic Y, Coe IR (2002) Molecular evolution of the equilibrative nucleoside transporter family: identification of novel family members in prokaryotes and eukaryotes. Mol Biol Evol 19:2199–2210

    Article  CAS  PubMed  Google Scholar 

  • Akerman M, Mandel-Gutfreund Y (2006) Alternative splicing regulation at tandem 3ʹ splice sites. Nucleic Acids Res 34:23–31

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bernard C, Traub M, Kunz HH, Hach S, Trentmann O, Mohlmann T (2011) Equilibrative nucleoside transporter 1 (ENT1) is critical for pollen germination and vegetative growth in Arabidopsis. J Exp Bot 62:4627–4637

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Campbell MA, Haas BJ, Hamilton JP, Mount SM, Buell CR (2006) Comprehensive analysis of alternative splicing in rice and comparative analyses with Arabidopsis. BMC Genomics 7:327

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Cao SQ, Jiang L, Song SY, Jing R, Xu GS (2006) AtGRP7 is involved in the regulation of abscisic acid and stress responses in Arabidopsis. Cell Mol Biol Lett 11:526–535

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Cheng Q, Zhou Y, Liu Z, Zhang L, Song G, Guo Z, Wang W, Qu X, Zhu Y, Yang D (2015) An alternatively spliced heat shock transcription factor, OsHSFA2dI, functions in the heat stress-induced unfolded protein response in rice. Plant Biol 17:419–429

    Article  CAS  PubMed  Google Scholar 

  • Chern TM, van Nimwegen E, Kai C, Kawai J, Carninci P, Hayashizaki Y, Zavolan M (2006) A simple physical model predicts small exon length variations. PLoS Genet 2:606–613

    Article  CAS  Google Scholar 

  • Darracq A, Adams KL (2013) Features of evolutionarily conserved alternative splicing events between Brassica and Arabidopsis. New Phytol 199:252–263

    Article  CAS  PubMed  Google Scholar 

  • Dinesh-Kumar SP, Baker BJ (2000) Alternatively spliced N resistance gene transcripts: their possible role in tobacco mosaic virus resistance. Proc Natl Acad Sci USA 97:1908–1913

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Filichkin SA, Priest HD, Givan SA, Shen R, Bryant DW, Fox SE, Wong WK, Mockler TC (2010) Genome-wide mapping of alternative splicing in Arabidopsis thaliana. Genome Res 20:45–58

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Filichkin S, Priest HD, Megraw M, Mockler TC (2015) Alternative splicing in plants: directing traffic at the crossroads of adaptation and environmental stress. Curr Opin Plant Biol 24:125–135

    Article  CAS  PubMed  Google Scholar 

  • Fu ZQ, Guo M, Jeong BR, Tian F, Elthon TE, Cerny RL, Staiger D, Alfano JR (2007) A type III effector ADP-ribosylates RNA-binding proteins and quells plant immunity. Nature 447:284–288

    Article  CAS  PubMed  Google Scholar 

  • Iniguez LP, Ramirez M, Barbazuk WB, Hernandez G (2017) Identification and analysis of alternative splicing events in Phaseolus vulgaris and Glycine max. BMC Genomics 18:650

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Jackson S, Chen ZJ (2010) Genomic and expression plasticity of polyploidy. Curr Opin Plant Biol 13:153–159

    Article  CAS  PubMed  Google Scholar 

  • Jiang JJ, Wang Y, Zhu B, Fang TT, Fang YJ, Wang YP (2015) Digital gene expression analysis of gene expression differences within Brassica diploids and allopolyploids. BMC Plant Biol 15:22

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Keller M, Hu YJ, Mesihovic A, Fragkostefanakis S, Schleiff E, Simm S (2017) Alternative splicing in tomato pollen in response to heat stress. DNA Res 24:205–217

    Article  CAS  PubMed  Google Scholar 

  • Ko DK, Rohozinski D, Song QX, Taylor SH, Juenger TE, Harmon FG, Chen ZJ (2016) Temporal shift of circadian-mediated gene expression and carbon fixation contributes to biomass heterosis in maize hybrids. PLoS Genet 12:e1006197

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Li B, Dewey CN (2011) RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome. BMC Bioinformatics 12:323

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Li H, Ruan J, Durbin R (2008) Mapping short DNA sequencing reads and calling variants using mapping quality scores. Genome Res 18:1851–1858

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Li AL, Liu DC, Wu J, Zhao XB, Hao M, Geng SF, Yan J, Jiang XX, Zhang LQ, Wu JY, Yin LJ, Zhang RZ, Wu L, Zheng YL, Mao L (2014) MRNA and small RNA transcriptomes reveal insights into dynamic homoeolog regulation of allopolyploid heterosis in nascent hexaploid wheat. Plant Cell 26:1878–1900

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Liu ZS, Qin JX, Tian XJ, Xu SB, Wang Y, Li HX, Wang XM, Peng HR, Yao YY, Hu ZR, Ni ZF, Xin MM, Sun QX (2018) Global profiling of alternative splicing landscape responsive to drought, heat and their combination in wheat (Triticum asetivum L.). Plant Biotechnol J 16:714–726

    Article  CAS  PubMed  Google Scholar 

  • Lorkovic ZJ, Lehner R, Forstner C, Barta A (2005) Evolutionary conservation of minor U12-type spliceosome between plants and humans. RNA 11:1095–1107

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lu TT, Lu GJ, Fan DL, Zhu CR, Li W, Zhao QA, Feng Q, Zhao Y, Guo YL, Li WJ, Huang XH, Han B (2010) Function annotation of the rice transcriptome at single-nucleotide resolution by RNA-seq. Genome Res 20:1238–1249

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mandadi KK, Scholthof KB (2015) Genome-wide analysis of alternative splicing landscapes modulated during plant-virus interactions in Brachypodium distachyon. Plant Cell 27:71–85

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Marquez Y, Brown JWS, Simpson C, Barta A, Kalyna M (2012) Transcriptome survey reveals increased complexity of the alternative splicing landscape in Arabidopsis. Genome Res 22:1184–1195

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mei WB, Liu SZ, Schnable JC, Yeh CT, Springer NM, Schnable PS, Barbazuk WB (2017) A comprehensive analysis of alternative splicing in paleopolyploid maize. Front Plant Sci 8:694

    Article  PubMed  PubMed Central  Google Scholar 

  • Ottens F, Gehring NH (2016) Physiological and pathophysiological role of nonsense-mediated mRNA decay. Pflug Arch Eur J Phy 468:1013–1028

    Article  CAS  Google Scholar 

  • Qi B, Huang W, Zhu B, Zhong XF, Guo JH, Zhao N, Xu CM, Zhang HK, Pang JS, Han FP, Liu B (2012) Global transgenerational gene expression dynamics in two newly synthesized allohexaploid wheat (Triticum aestivum) lines. BMC Biol 10:3

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Romanowski A, Yanovsky MJ (2015) Circadian rhythms and post-transcriptional regulation in higher plants. Front Plant Sci 6:437

    Article  PubMed  PubMed Central  Google Scholar 

  • Saminathan T, Nimmakayala P, Manohar S et al (2014) Differential gene expression and alternative splicing between diploid and tetraploid watermelon. J Exp Bot 66:1369–1385

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Schmal C, Reimann P, Staiger D (2013) A circadian clock-regulated toggle switch explains AtGRP7 and AtGRP8 oscillations in Arabidopsis thaliana. PLoS Comput Biol 9:e1002986

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Schmidt F, Marnef A, Cheung MK, Wilson I, Hancock J, Staiger D, Ladomery M (2010) A proteomic analysis of oligo(dT)-bound mRNP containing oxidative stress-induced Arabidopsis thaliana RNA-binding proteins ATGRP7 and ATGRP8. Mol Biol Rep 37:839–845

    Article  CAS  PubMed  Google Scholar 

  • Schoning JC, Streitner C, Meyer IM, Gao YH, Staiger D (2008) Reciprocal regulation of glycine-rich RNA-binding proteins via an interlocked feedback loop coupling alternative splicing to nonsense-mediated decay in Arabidopsis. Nucleic Acids Res 36:6977–6987

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Seo PJ, Park MJ, Park CM (2013) Alternative splicing of transcription factors in plant responses to low temperature stress: mechanisms and functions. Planta 237:1415–1424

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shen SH, Park JW, Lu ZX, Lin L, Henry MD, Wu YN, Zhou Q, Xing Y (2014a) rMATS: robust and flexible detection of differential alternative splicing from replicate RNA-Seq data. Proc Natl Acad Sci USA 111:E5593–E5601

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shen YT, Zhou ZK, Wang Z, Li WY, Fang C, Wu M, Ma YM, Liu TF, Kong LA, Peng DL, Tian ZX (2014b) Global dissection of alternative splicing in paleopolyploid soybean. Plant Cell 26:996–1008

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shin KH, Yang SH, Lee JY, Lim CW, Lee SC, Brown JW, Kim SH (2015) Alternative splicing of mini-exons in the Arabidopsis leaf rust receptor-like kinase LRK10 genes affects subcellular localization. Plant Cell Rep 34:495–505

    Article  CAS  PubMed  Google Scholar 

  • Soltis DE, Visger CJ, Marchant DB, Soltis PS (2016) Polyploidy: pitfalls and paths to a paradigm. Am J Bot 103:1146–1166

    Article  PubMed  Google Scholar 

  • Song KM, Lu P, Tang K, Osborn TC (1995) Rapid genome change in synthetic polyploids of Brassica and its implications for polyploid evolution. Proc Natl Acad Sci USA 92:7719–7723

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Staiger D, Zecca L, Kirk DAW, Apel K, Eckstein L (2003) The circadian clock regulated RNA-binding protein AtGRP7 autoregulates its expression by influencing alternative splicing of its own pre-mRNA. Plant J 33:361–371

    Article  CAS  PubMed  Google Scholar 

  • Streitner C, Danisman S, Wehrle F, Schoning JC, Alfano JR, Staiger D (2008) The small glycinerich RNA binding protein AtGRP7 promotes floral transition in Arabidopsis thaliana. Plant J 56:239–250

    Article  CAS  PubMed  Google Scholar 

  • Szecsi J, Joly C, Bordji K, Varaud E, Cock JM, Dumas C, Bendahmane M (2006) BIGPETALp, a bHLH transcription factor is involved in the control of Arabidopsis petal size. EMBO J 25:3912–3920

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Tack DC, Pitchers WR, Adams KL (2014) Transcriptome analysis indicates considerable divergence in alternative splicing between duplicated genes in Arabidopsis thaliana. Genetics 198:473

    Article  CAS  Google Scholar 

  • Tadokoro K, Yamazaki-Inoue M, Tachibana M, Fujishiro M, Nagao K, Toyoda M, Ozaki M, Ono M, Miki N, Miyashita T, Yamada M (2005) Frequent occurrence of protein isoforms with or without a single amino acid residue by subtle alternative splicing: the case of Gln in DRPLA affects subcellular localization of the products. Am J Hum Genet 50:382–394

    Article  CAS  Google Scholar 

  • Tarazona S, Garcia-Alcalde F, Dopazo J, Ferrer A, Conesa A (2011) Differential expression in RNA-seq: a matter of depth. Genome Res 21:2213–2223

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Terashima A, Takumi S (2009) Allopolyploidization reduces alternative splicing efficiency for transcripts of the wheat DREB2 homolog, WDREB2. Genome 52:100–105

    Article  CAS  PubMed  Google Scholar 

  • Tian ET, Jiang YF, Chen LL, Zou J, Liu F, Meng JL (2010) Synthesis of a Brassica trigenomic allohexaploid (B. carinata × B. rapa) de novo and its stability in subsequent generations. Theor Appl Genet 121:1431–1440

    Article  CAS  PubMed  Google Scholar 

  • Trapnell C, Pachter L, Salzberg SL (2009) TopHat: discovering splice junctions with RNA-Seq. Bioinformatics 25:1105–1111

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Trapnell C, Williams BA, Pertea G, Mortazavi A, Kwan G, van Baren MJ, Salzberg SL, Wold BJ, Pachter L (2010) Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat Biotechnol 28:511–1174

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • van Nocker S, Vierstra RD (1993) Two cDNAs from Arabidopsis thaliana encode putative RNA binding proteins containing glycine-rich domain. Plant Mol Biol 21:695–699

    Article  PubMed  Google Scholar 

  • Wang JL, Tian L, Lee HS, Wei NE, Jiang HM, Watson B, Madlung A, Osborn TC, Doerge RW, Comai L, Chen ZJ (2006) Genome-wide nonadditive gene regulation in Arabidopsis allotetraploids. Genetics 172:507–517

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wang MJ, Wang PC, Liang F, Ye ZX, Li JY, Shen C, Pei LL, Wang F, Hu J, Tu LL, Lindsey K, He DH, Zhang XL (2017) A global survey of alternative splicing in allopolyploid cotton: landscape, complexity and regulation. New Phytol 217:163–178

    Article  CAS  PubMed  Google Scholar 

  • Wang Z, Zhang H, Gong W (2019) Genome-wide identification and comparative analysis of alternative splicing across four legume species. Planta 249:1133–1142

    Article  CAS  PubMed  Google Scholar 

  • Zhang XC, Gassmann W (2003) RPS4-mediated disease resistance requires the combined presence of RPS4 transcripts with full length and truncated open reading frames. Plant Cell 15:2333–2342

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhang ZG, Zhou L, Wang P, Liu Y, Chen XF, Hu LDA, Kong XY (2009) Divergence of exonic splicing elements after gene duplication and the impact on gene structures. Genome Biol 10:R120

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhang GJ, Guo GW, Hu XD et al (2010a) Deep RNA sequencing at single base-pair resolution reveals high complexity of the rice transcriptome. Genome Res 20:646–654

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhang PG, Huang SZ, Pin AL, Adams KL (2010b) Extensive divergence in alternative splicing patterns after gene and genome duplication during the evolutionary history of Arabidopsis. Mol Biol Evol 27:1686–1697

    Article  CAS  PubMed  Google Scholar 

  • Zhou R, Moshgabadi N, Adams KL (2011) Extensive changes to alternative splicing patterns following allopolyploidy in natural and resynthesized polyploids. Proc Natl Acad Sci USA 108:16122–16127

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This work was supported by the National Natural Science Foundation of China (31570539, 31370258).

Author information

Authors and Affiliations

  1. State Key Laboratory of Hybrid Rice, College of Life Sciences, Wuhan University, Wuhan, China

    Ruihua Wang, Helian Liu, Zhengyi Liu & Jianbo Wang

  2. National Key Laboratory of Crop Genetic Improvement, College of Plant Science and Technology, Huazhong Agricultural University, Wuhan, China

    Jun Zou & Jinling Meng

Authors
  1. Ruihua Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Helian Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Zhengyi Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jun Zou
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Jinling Meng
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Jianbo Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jianbo Wang.

Ethics declarations

Conflict of interest

The authors declare that they have no conflicts of interest.

Availability of data and materials

The raw data of RNA-seq reads were deposited in the NCBI database under accession number (SRR6936350, SRR6936349, SRR6936348, SRR6936347, SRR6936346, SRR6936345, SRR6936344, SRR6936343, SRR6936342).

Additional information

Publisher's Note

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

Electronic supplementary material

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Wang, R., Liu, H., Liu, Z. et al. Genome-wide analysis of alternative splicing divergences between Brassica hexaploid and its parents. Planta 250, 603–628 (2019). https://doi.org/10.1007/s00425-019-03198-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00425-019-03198-z

Keywords

Search

Navigation