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Recessive male sterility in cabbage (Brassica oleracea var. capitata) caused by loss of function of BoCYP704B1 due to the insertion of a LTR-retrotransposon

Abstract

Key message

The LTR-retrotransposon insertion in BoCYP704B1 is proved to be the primary cause of the male sterility in cabbage. Effective allele-specific markers were developed for marker-assisted selection of male sterile gene.

Abstract

83121A is a spontaneous male sterile mutant identified from cabbage. Genetic analysis indicated that male sterility is controlled by a single recessive gene. Pollen wall formation in the 83121A mutant was severely defective, with a lack of sporopollenin or exine. To understand the mechanisms of male sterility in 83121A, transcription analysis using RNA-Seq was carried out in the buds of the male sterile line 83121A and the male fertile line 83121B, which are near-isogenic lines differing only in the fertility trait. Via expression analysis of differentially expressed genes involved in pollen exine development before the bicellular pollen stage, BoCYP704B1 was identified as a candidate gene, which was approximately downregulated 30-fold in 83121A. BoCYP704B1 is a member of the evolutionarily conserved CYP704B family, which is essential for sporopollenin formation. The BoCYP704B1 transcript is specifically detected in the developing anthers of wild-type cabbage. Further sequence analysis revealed that a 5424-bp long terminal repeat-retrotransposon (LTR-RT) was inserted into the first exon of BoCYP704B1 in 83121A, which is not found in wild-type plants. The insertion of LTR-RT not only reduced the expression of BoCYP704B1 but also altered structure of protein encoded by BoCYP704B1. Moreover, linkage analysis showed that the homozygotic mutational BoCYP704B1 always cosegregated with male sterility. These data suggest that the LTR-RT insertion in BoCYP704B1 hinders sporopollenin formation in 83121A leading to male sterility. The allele-specific markers developed in this study were effective for marker-assisted selection of the male sterile gene.

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References

  • Ariizumi T, Toriyama K (2011) Genetic regulation of sporopollenin synthesis and pollen exine development. Annu Rev Plant Biol 62:437–460

    CAS  Article  PubMed  Google Scholar 

  • Barghini E, Natali L, Giordani T, Cossu RM, Scalabrin S, Cattonaro F, Šimková H, Vrána J, Doležel J, Morgante M, Cavallini A (2014) LTR retrotransposon dynamics in the evolution of the olive (Olea europaea) genome. DNA Res 22(1):91–100

    Article  PubMed  PubMed Central  Google Scholar 

  • Blackmore S, Wortley AH, Skvarla JJ, Rowley JR (2007) Pollen wall development in flowering plants. New Phytol 174(3):483–498

    CAS  Article  PubMed  Google Scholar 

  • Butelli E, Licciardello C, Zhang Y, Liu J, Mackay S, Bailey P, Reforgiato-Recupero G, Martin C (2012) Retrotransposons control fruit-specific, cold-dependent accumulation of anthocyanins in blood oranges. Plant Cell 24(3):1242–1255

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Chen W, Yu X, Zhang K, Shi J, De Oliveira S, Schreiber L, Shanklin J, Zhang D (2011) Male Sterile2 encodes a plastid-localized fatty acyl carrier protein reductase required for pollen exine development in Arabidopsis. Plant Physiol 157(2):842–853

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Conesa A, Götz S (2008) Blast2GO: a comprehensive suite for functional analysis in plant genomics. Int J Plant Genom 2008:619832

    Google Scholar 

  • Dayhoff M (1977) A model of evolutionary change in proteins. Atlas Protein Seq Struct 5:345–352

    Google Scholar 

  • de Azevedo Souza C, Kim SS, Koch S, Kienow L, Schneider K, McKim SM, Haughn GW, Kombrink E, Douglas CJ (2009) A novel fatty acyl-CoA synthetase is required for pollen development and sporopollenin biosynthesis in Arabidopsis. Plant Cell 21(2):507–525

    Article  Google Scholar 

  • Dobritsa AA, Shrestha J, Morant M, Pinot F, Matsuno M, Swanson R, Møller BL, Preuss D (2009) Cyp704b1 is a long-chain fatty acid ω-hydroxylase essential for sporopollenin synthesis in pollen of arabidopsis. Plant Physiol 151(2):574–589

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Dobritsa AA, Lei Z, Nishikawa S, Urbanczyk-Wochniak E, Huhman DV, Preuss D, Sumner LW (2010) LAP5 and LAP6 encode anther-Specific proteins with similarity to chalcone synthase essential for pollen exine development in arabidopsis. Plant physiol 153(3):937–955

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Doyle JJ (1990) Isolation of plant DNA from fresh tissue. Focus 12:13–15

    Google Scholar 

  • Du J, Tian Z, Hans CS, Laten HM, Cannon SB, Jackson SA, Shoemaker RC, Ma J (2010) Evolutionary conservation, diversity and specificity of LTR-retrotransposons in flowering plants: insights from genome-wide analysis and multi-specific comparison. Plant J 63(4):584–598

    CAS  Article  PubMed  Google Scholar 

  • Grienenberger E, Kim SS, Lallemand B, Geoffroy P, Heintz D, de Azevedo Souza C, Heitz T, Douglas CJ, Legrand M (2010) Analysis of TETRAKETIDE α-PYRONE REDUCTASE function in Arabidopsis thaliana reveals a previously unknown, but conserved, biochemical pathway in sporopollenin monomer biosynthesis. Plant Cell 22(12):4067–4083

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Guo AY, Zhu QH, Chen X, Luo JC (2007) GSDS: a gene structure display server. Hereditas/Zhongguo yi chuan xue hui bian ji=Yi Chuan 29(8):1023–1026

    CAS  Article  Google Scholar 

  • Hudakova S, Michalek W, Presting GG, ten Hoopen R, dos Santos K, Jasencakova Z, Schubert I (2001) Sequence organization of barley centromeres. Nucl Acids Res 29(24):5029–5035

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Jiang J, Zhang Z, Cao J (2013) Pollen wall development: the associated enzymes and metabolic pathways. Plant Biol 15(2):249–263

    CAS  Article  PubMed  Google Scholar 

  • Kanehisa M, Kanehisa M, Araki M, Goto S, Hattori M, Hirakawa M, Itoh M, Katayama T, Kawashima S, Okuda S, Tokimatsu T, Yamanishi Y (2008) KEGG for linking genomes to life and the environment. Nucl Acids Res 36(suppl 1):D480–D484

    CAS  PubMed  Google Scholar 

  • Katoh K, Standley DM (2013) MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol 30(4):772–780

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Kim SS, Grienenberger E, Lallemand B, Colpitts CC, Kim SY, de Azevedo Souza C, Geoffroy P, Heintz D, Krahn D, Kaiser M, Kombrink E, Heitz T, Suh D, Legrand M, Douglas CJ (2010) LAP6/POLYKETIDE SYNTHASE A and LAP5/POLYKETIDE SYNTHASE B encode hydroxyalkyl α-pyrone synthases required for pollen development and sporopollenin biosynthesis in Arabidopsis thaliana. The Plant Cell 22(12):4045–4066

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Kobayashi S, Goto-Yamamoto N, Hirochika H (2004) Retrotransposon-induced mutations in grape skin color. Science 304(5673):982–982

    Article  PubMed  Google Scholar 

  • Kumar A, Bennetzen JL (1999) Plant retrotransposons. Annu Rev Genet 33(1):479–532

    CAS  Article  PubMed  Google Scholar 

  • Kumekawa N, Ohmido N, Fukui K, Ohtsubo E, Ohtsubo H (2001) A new gypsy-type retrotransposon, RIRE7: preferential insertion into the tandem repeat sequence TrsD in pericentromeric heterochromatin regions of rice chromosomes. Mol Gen Genomics 265(3):480–488

    CAS  Article  Google Scholar 

  • Langmead B, Trapnell C, Pop M, Salzberg SL (2009) Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol 10(3):1

    Article  Google Scholar 

  • Li H, Pinot F, Sauveplane V, Werck-Reichhart D, Diehl P, Schreiber L, Franke R, Zhang P, Chen L, Gao Y, Liang W, Zhang D (2010) Cytochrome p450 family member cyp704b2 catalyzes the {omega}-hydroxylation of fatty acids and is required for anther cutin biosynthesis and pollen exine formation in rice. Plant Cell 22(1):173–190

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Lisch D (2013) How important are transposons for plant evolution? Nat Rev Genet 14(1):49–61

    CAS  Article  PubMed  Google Scholar 

  • Liu L, Fan XD (2013) Tapetum: regulation and role in sporopollenin biosynthesis in Arabidopsis. Plant Mol Biol 83(3):165–175

    CAS  Article  PubMed  Google Scholar 

  • Liu X, Wu J, Wang J, Liu X, Zhao S, Li Z, Kong L, Gu X, Luo J, Gao G (2009) Weblab: a data-centric, knowledge-sharing bioinformatic platform. Nucl Acids Research 37(Web Server issue):33–39

    Google Scholar 

  • Liu S, Liu Y, Yang X et al (2014) The Brassica oleracea genome reveals the asymmetrical evolution of polyploid genomes. Nat Commun 5:3930

    CAS  PubMed  PubMed Central  Google Scholar 

  • Lou P, Kang J, Zhang G, Bonnema G, Fang Z, Wang X (2007) Transcript profiling of a dominant male sterile mutant (Ms-cd1) in cabbage during flower bud development. Plant Sci 172(1):111–119

    CAS  Article  Google Scholar 

  • Ma Y, Kang J, Wu J, Zhu Y, Wang X (2015) Identification of tapetum-specific genes by comparing global gene expression of four different male sterile lines in Brassica oleracea. Plant Mol Biol 87(6):541–554

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Morant M, Jørgensen K, Schaller H, Pinot F, Møller BL, Werck-Reichhart D, Bak S (2007) CYP703 is an ancient cytochrome P450 in land plants catalyzing in-chain hydroxylation of lauric acid to provide building blocks for sporopollenin synthesis in pollen. Plant Cell 19(5):1473–1487

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Mortazavi A, Williams BA, McCue K, Schaeffer L, Wold B (2008) Mapping and quantifying mammalian transcriptomes by RNA-SEq. Nat Methods 5(7):621–628

    CAS  Article  PubMed  Google Scholar 

  • Perez-Prat E, van Lookeren Campagne MM (2002) Hybrid seed production and the challenge of propagating male-sterile plants. Trends Plant Sci 7(5):199–203

    CAS  Article  PubMed  Google Scholar 

  • Quilichini TD, Samuels AL, Douglas CJ (2014) ABCG26-mediated polyketide trafficking and hydroxycinnamoyl spermidines contribute to pollen wall exine formation in Arabidopsis. Plant Cell 26(11):4483–4498

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Ronquist F, Teslenko M, Van Der Mark P, Ayres DL, Darling A, Höhna S, Larget B, Liu L, Suchard MA, Huelsenbeck JP (2012) MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Syst Biol 61(3):539–542

    Article  PubMed  PubMed Central  Google Scholar 

  • Sivaguru M, Mander L, Fried G, Punyasena SW (2012) Capturing the surface texture and shape of pollen: a comparison of microscopy techniques. PloS One 7(6):e39129

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Tian Z, Rizzon C, Du J, Zhu L, Bennetzen JL, Jackson SA, Gaut BS, Ma J (2009) Do genetic recombination and gene density shape the pattern of DNA elimination in rice long terminal repeat retrotransposons? Genome Res 19(12):2221–2230

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Trapnell C, Pachter L, Salzberg SL (2009) TopHat: discovering splice junctions with RNA-SEq. Bioinformatics 25(9):1105–1111

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  • Vedel F, Pla M, Vitart V, Gutierres S, Chetrit P, De Paepe R (1994) Molecular basis of nuclear and cytoplasmic male sterility in higher plants. Plant Physiol BioChem 32(5):601–618

    CAS  Google Scholar 

  • Xiong Y, Eickbush TH (1990) Origin and evolution of retroelements based upon their reverse transcriptase sequences. EMBO J 9(10):3353

    CAS  PubMed  PubMed Central  Google Scholar 

  • Yang M, Zhu L, Pan C, Xu L, Liu Y, Ke W, Yang P (2015) Transcriptomic analysis of the regulation of rhizome formation in temperate and tropical lotus (Nelumbo nucifera). Sci Rep 5(4):180–181

    Google Scholar 

  • Yi B, Zeng F, Lei S, Chen Y, Yao X, Zhu Y, Wen J, Shen J, Ma C, Tu J, Fu T (2010) Two duplicate CYP704B1-homologous genes BnMs1 and BnMs2 are required for pollen exine formation and tapetal development in Brassica napus. Plant J 63(6):925–938

    CAS  Article  PubMed  Google Scholar 

  • Zhang L, Mao D, Xing F, Bai X, Zhao H, Yao W, Li G, Xie W, Xing Y (2015) Loss of function of OsMADS3 via the insertion of a novel retrotransposon leads to recessive male sterility in rice (Oryza sativa). Plant Sci 238:188–197

    CAS  Article  PubMed  Google Scholar 

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Acknowledgements

This work was funded by the National Key Research and Development Program (SQ2017ZY030004), the Science and Technology Innovation Program of the Chinese Academy of Agricultural Sciences (CAAS-ASTIP-2013-IVFCAAS), the Modern Agro-Industry Technology Research System (CARS-25-B-01), the National High Technology Research and Development Program of China (863 Program, 2012AA100101), the Key Projects in the National Science and Technology Pillar Program during the Twelfth Five-Year Plan Period (2012BAD02B01), and the Project of Science and Technology Commission of Beijing Municipality (Z141105002314020-1). The authors thank LetPub (http://www.letpub.com) for its linguistic assistance during the preparation of this manuscript.

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Correspondence to Li-mei Yang.

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Communicated by Maria Laura Federico.

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122_2017_2899_MOESM1_ESM.tif

Supplementary Fig. S1 SEM micrographs of pollen from 83121A plants (a, c) and 83121B plants (b, d). Scale bars = 10 μm (a, b) and 2 μm (TIF 643 KB)

Supplementary Fig. S2 Correlation analysis of gene expression levels between replicate samples (TIF 1840 KB)

122_2017_2899_MOESM3_ESM.tif

Supplementary Fig. S3 Protein phylogeny of the CYP704 family and related P450 enzymes. Phylogenetic analysis was performed using MrBayes v3.2.1 based on the Dayhoff model for 100,000 generations. At, Arabidopsis; Bn, Brassica napus; Mt, Medicago truncatula; Os, rice; Pp, Physcomitrella patens; Pta, Pinus taeda; Ptr, Populus trichocarpa; Sm, Selaginella moellendorffii; Vv, Vitis vinifera; Zm, Zea mays (TIF 532 KB)

122_2017_2899_MOESM4_ESM.tif

Supplementary Fig. S4 Predicted proteins encoded by BoCYP704B1 in wild-type and 83121A. BoCYP704B1 sequence in 83121A is completely changed with the exception of the fragment in black box (TIF 447 KB)

Supplementary Table 1 (DOCX 12 KB)

Supplementary Table 2 (DOCX 14 KB)

Supplementary Table 3 (DOCX 16 KB)

Supplementary Table 4 (DOCX 12 KB)

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Ji, Jl., Yang, Lm., Fang, Zy. et al. Recessive male sterility in cabbage (Brassica oleracea var. capitata) caused by loss of function of BoCYP704B1 due to the insertion of a LTR-retrotransposon. Theor Appl Genet 130, 1441–1451 (2017). https://doi.org/10.1007/s00122-017-2899-z

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Keywords

  • Male Sterility
  • Cytoplasmic Male Sterility
  • Restorer Gene
  • Male Sterile Plant
  • 83121A Mutant