Skip to main content
SpringerLink
Log in

Isolation and Characterization of an Anther-Specific Polygalacturonase Gene, BcMF16, in Brassica campestris ssp. chinensis

  • Published:
Plant Molecular Biology Reporter Aims and scope Submit manuscript

Abstract

Many genes in the genic male sterile A/B line (Bajh97-01A/B) of Chinese cabbage pak choi (Brassica campestris L. subsp. chinensis Makino) are expressed differentially, and some play critical roles in the formation of pollen walls. In this study, one of these genes, Brassica campestris Male Fertility 16 (BcMF16), has been isolated and characterized. The BcMF16 gene shares approximately 85% nucleotide sequence homology with two exopolygalacturonase (EC3.2.1.67) genes of Arabidopsis thaliana. Cluster analysis of polygalacturonase peptides indicate that BcMF16 belongs to the pollen polygalacturonase clade. Quantitative real-time PCR analysis has revealed that BcMF16 is specifically expressed in reproductive tissues of the fertile line of genic male sterile A/B line of Chinese cabbage pak choi, and that expression levels dramatically increased during later stages of pollen development. In situ hybridization has demonstrated that BcMF16 is specifically and transiently expressed in both tapetum and pollen following microspore separation at the tetrad stage.

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

Similar content being viewed by others

Abbreviations

cDNA-AFLP:

cDNA-amplified fragment length polymorphism

ORF:

Open reading frame

PG:

Polygalacturonase

RACE:

Rapid amplification of cDNA ends

RT-PCR:

Reverse transcriptase polymerase chain reaction

BcMF:

Brassica campastris Male Fertile

References

  • Bairoch A, Bucher P, Hofmann K (1997) The PROSITE database, its status in 1997. Nucleic Acids Res 25:217–221

    Article  PubMed  CAS  Google Scholar 

  • Bendtsen JD, Nielsen H, von Heijne G, Brunak S (2004) Improved prediction of signal peptides: SignalP 3.0. J Mol Biol 340:783–795

    Article  PubMed  Google Scholar 

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

    Article  PubMed  CAS  Google Scholar 

  • Chen W, Baldwin TC (2007) An improved method for the fixation, embedding and immunofluorescence labeling of resin-embedded plant tissue. Plant Mol Biol Rep 25:27–35

    Article  CAS  Google Scholar 

  • Chen WJ, Chang SH, Hudson ME, Kwan WK, Li J, Estes B, Knoll D, Shi L, Zhu T (2005) Contribution of transcriptional regulation to natural variations in Arabidopsis. Genome Biol 6:R32

    Article  PubMed  Google Scholar 

  • Deuel H, Stutz E (1958) Pectic substances and pectic enzymes. Adv Enzymol Relat Subj Biochem 20:341–382

    PubMed  CAS  Google Scholar 

  • Drakakaki G, Zabotina O, Delgado I, Robert S, Keegstra K, Raikhel N (2006) Arabidopsis reversibly glycosylated polypeptides 1 and 2 are essential for pollen development. Plant Physiol 142:1480–1492

    Article  PubMed  CAS  Google Scholar 

  • Francis KE, Lam SY, Copenhaver GP (2006) Separation of Arabidopsis pollen tetrads is regulated by QUARTET1, a pectin methylesterase gene. Plant Physiol 142:1004–1013

    Article  PubMed  CAS  Google Scholar 

  • Goda H, Sawa S, Asami T, Fujioka S, Shimada Y, Yoshida S (2004) Comprehensive comparison brassinosteroid-regulated of auxin-regulated and brassinosteroid-regulated genes in Arabidopsis. Plant Physiol 134:1555–1573

    Article  PubMed  CAS  Google Scholar 

  • Guyon VN, Astwood JD, Garner EC, Dunker AK, Taylor LP (2000) Isolation and characterization of cDNAs expressed in the early stages of flavonol-induced pollen germination in petunia. Plant Physiol 123:699–710

    Article  PubMed  CAS  Google Scholar 

  • Hadfield KA, Bennett AB (1998) Polygalacturonases: many genes in search of a function. Plant Physiol 117:337–343

    Article  PubMed  CAS  Google Scholar 

  • Hasegawa S, Nagel CW (1968) Isolation of an oligogalacturonate hydrolase from a Bacillus specie. Arch of Biochem and Biophys 124:513–520

    Article  CAS  Google Scholar 

  • Hiller K, Grote A, Scheer M, Munch R, Jahn D (2004) PrediSi: prediction of signal peptides and their cleavage positions. Nucleic Acids Res 32:W375–379

    Article  PubMed  CAS  Google Scholar 

  • Holmes-Davis R, Tanaka CK, Vensel WH, Hurkman WJ, McCormick S (2005) Proteome mapping of mature pollen of Arabidopsis thaliana. Proteomics 5:4864–4884

    Article  PubMed  CAS  Google Scholar 

  • Honys D, Twell D (2003) Comparative analysis of the Arabidopsis pollen transcriptome. Plant Physiol 132:640–652

    Article  PubMed  CAS  Google Scholar 

  • Honys D, Oh SA, Solcova B, Johnson JA, Boudova R, Douda J, Twell D (2005) New tools for the manipulation of microspore gene expression. Acta Biol Cracov Bot 47:58–58

    Google Scholar 

  • Huang L, Cao J, Ye W, Liu T, Jiang L, Ye Y (2008) Transcriptional differences between the male-sterile mutant bcms and wild-type Brassica campestris ssp. chinensis reveal genes related to pollen development. Plant Biol 10:342–355

    Article  PubMed  CAS  Google Scholar 

  • Huang L, Cao JS, Zhang AH, Ye YQ, Zhang YC, Liu TT (2009a) The polygalacturonase gene BcMF2 from Brassica campestris is associated with intine development. J Exp Bot 60:301–313

    Article  PubMed  CAS  Google Scholar 

  • Huang L, Ye Y, Zhang Y, Zhang A, Liu T, Cao J (2009b) BcMF9, a novel polygalacturonase gene, is required for both Brassica campestris intine and exine formation. Ann Bot 104:1339–1351

    Article  PubMed  CAS  Google Scholar 

  • Jones-Rhoades MW, Borevitz JO, Preuss D (2007) Genome-wide expression profiling of the Arabidopsis female gametophyte identifies families of small, secreted proteins. PLoS Genet 3:1848–1861

    Article  PubMed  CAS  Google Scholar 

  • Kato H, Xie GS, Sato Y, Imai R (2010) Isolation of anther-specific gene promoters suitable for transgene expression in rice. Plant Mol Biol Rep 28:381–387

    Article  CAS  Google Scholar 

  • Kehr J, Buhtz A, Giavalisco P (2005) Analysis of xylem sap proteins from Brassica napus. BMC Plant Biol 5:11

    Article  PubMed  Google Scholar 

  • Kim J, Patterson SE (2006) Expression divergence and functional redundancy of polygalacturonases in floral organ abscission. Plant Signal Behav 1:281–283

    Article  PubMed  Google Scholar 

  • Lau JM, Cooper NG, Robinson DL, Korban SS (2009) Sequence and in silico characterization of the tomato polygalacturonase (PG) promoter and terminator regions. Plant Mol Biol Rep 27:250–256

    Article  CAS  Google Scholar 

  • Liu YZ, Dong T, Lei Y, Deng XX, Gu QQ (2011) Isolation of a polygalacturonase gene from Citrus sinensis fruit and its expression relative to fruit mastication trait, fruit development, and calcium or boron treatments. Plant Mol Biol Rep 29:51–59

    Article  CAS  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) method. Methods 25:402–408

    Article  PubMed  CAS  Google Scholar 

  • Luo SN, Tai H, Zebarth B, Li XQ, Millard P, De Koeyer D, Xiong XY (2011) Sample collection protocol effects on quantification of gene expression in potato leaf tissue. Plant Mol Biol Rep 29:369–378

    Article  CAS  Google Scholar 

  • Malik MR, Wang F, Dirpaul JM, Zhou N, Polowick PL, Ferrie AM, Krochko JE (2007) Transcript profiling and identification of molecular markers for early microspore embryogenesis in Brassica napus. Plant Physiol 144:134–154

    Article  PubMed  CAS  Google Scholar 

  • Malik MR, Wang F, Dirpaul JM, Zhou N, Hammerlindl J, Keller W, Abrams SR, Ferrie AMR, Krochko JE (2008) Isolation of an embryogenic line from non-embryogenic Brassica napus cv. Westar through microspore embryogenesis. J Exp Bot 59:2857–2873

    Article  PubMed  CAS  Google Scholar 

  • Mc CR, Seegmiller CG (1954) Action of pectic enzymes on oligogalacturonic acids and some of their derivatives. Arch Biochem Biophys 50:440–450

    Article  Google Scholar 

  • Mill PJ (1966) The pectic enzymes of Aspergillus niger. A mercury-activated exopolygalacturonase. Biochem J 99:557–561

    PubMed  CAS  Google Scholar 

  • Morant M, Jorgensen K, Schaller H, Pinot F, Moller 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 19:1473–1487

    Article  PubMed  CAS  Google Scholar 

  • Nielsen H, Engelbrecht J, Brunak S, von Heijne G (1997) A neural network method for identification of prokaryotic and eukaryotic signal peptides and prediction of their cleavage sites. Int J Neural Syst 8:581–599

    Article  PubMed  CAS  Google Scholar 

  • Ogawa M, Kay P, Wilson S, Swain SM (2009) ARABIDOPSIS DEHISCENCE ZONE POLYGALACTURONASE1 (ADPG1), ADPG2, and QUARTET2 are polygalacturonases required for cell separation during reproductive development in Arabidopsis. Plant Cell 21:216–233

    Article  PubMed  CAS  Google Scholar 

  • Preuss D, Rhee SY, Davis RW (1994) Tetrad analysis possible in Arabidopsis with mutation of the QUARTET (QRT) genes. Science 264:1458–1460

    Article  PubMed  CAS  Google Scholar 

  • Qi JN, Yu SC, Zhang FL, Shen XQ, Zhao XY, Yu YJ, Zhang DS (2010) Reference gene selection for real-time quantitative polymerase chain reaction of mRNA transcript levels in Chinese cabbage (Brassica rapa L. ssp pekinensis). Plant Mol Biol Rep 28:597–604

    Article  CAS  Google Scholar 

  • Rhee SY, Somerville CR (1998) Tetrad pollen formation in quartet mutants of Arabidopsis thaliana is associated with persistence of pectic polysaccharides of the pollen mother cell wall. Plant J 15:79–88

    Article  PubMed  CAS  Google Scholar 

  • Rhee SY, Osborne E, Poindexter PD, Somerville CR (2003) Microspore separation in the quartet 3 mutants of Arabidopsis is impaired by a defect in a developmentally regulated polygalacturonase required for pollen mother cell wall degradation. Plant Physiol 133:1170–1180

    Article  PubMed  CAS  Google Scholar 

  • Rogstad SH, Keane B, Keiffer CH, Hebard F, Sisco P (2001) DNA extraction from plants: the use of pectinase. Plant Mol Biol Rep 19:353–359

    Article  CAS  Google Scholar 

  • Sitrit Y, Hadfield KA, Bennett AB, Bradford KJ, Downie AB (1999) Expression of a polygalacturonase associated with tomato seed germination. Plant Physiol 121:419–428

    Article  PubMed  CAS  Google Scholar 

  • vanDoorn WG, Stead AD (1997) Abscission of flowers and floral parts. J Exp Bot 48:821–837

    Article  CAS  Google Scholar 

  • Wang YQ, Yu XL, Cao JS (2004) Isolation and characterization of BcMF3, a gene expressed only in maintainer line in Chinese cabbage-pak-choi (Brassica campestris L. ssp. chinensis Makino var. communis Tsen et lee). J Genet Genomics 31:1302–1308

    Google Scholar 

  • Wang YQ, Ye WZ, Cao JS, Yu XL, Xiang X, Lu G (2005) Cloning and characterization of the microspore development-related gene BcMF2 in chinese cabbage pak-choi (Brassica campestris L. ssp. chinensis Makino). J Integr Plant Biol 47:9

    Google Scholar 

  • Wang Y, Zhang WZ, Song LF, Zou JJ, Su Z, Wu WH (2008) Transcriptome analyses show changes in gene expression to accompany pollen germination and tube growth in Arabidopsis. Plant Physiol 148:1201–1211

    Article  PubMed  CAS  Google Scholar 

  • Yang YZ, Singer SD, Liu ZR (2011) Petunia AGAMOUS enhancer-derived chimeric promoters specify a carpel-, stamen-, and petal-specific expression pattern sufficient for engineering male and female sterility in tobacco. Plant Mol Biol Rep 29:162–170

    Article  Google Scholar 

  • Zhang ZX, Tang WT, Zhang FD, Zheng YL (2005) Fertility restoration mechanisms in S-type cytoplasmic male sterility of maize (Zea mays L.) revealed through expression differences identified by cDNA microarray and suppression subtractive hybridization. Plant Mol Biol Rep 23:17–38

    Article  CAS  Google Scholar 

  • Zhang Q, Huang L, Liu TT, Yu XL, Cao JS (2008) Functional analysis of a pollen-expressed polygalacturonase gene BcMF6 in Chinese cabbage (Brassica campestris L. ssp chinensis Makino). Plant Cell Rep 27:1207–1215

    Article  PubMed  CAS  Google Scholar 

  • Zhu H, Yuan RC (2009) Abscisic acid, ethylene, and polygalacturonase are involved in young fruit abscission induced by NAA and shading in 'golden delicious' apples. Hortscience 44:1075–1075

    Google Scholar 

Download references

Acknowledgment

This work was supported by the Natural Science Foundation of China (no. 31071805) and the Natural Science Foundation of Zhejiang province (Y3090294).

Author information

Authors and Affiliations

  1. Laboratory of Cell & Molecular Biology, Institute of Vegetable Science, Zhejiang University, Hangzhou, 310029, People’s Republic of China

    Aihong Zhang, Lin Qiu, Li Huang, Xiaolin Yu, Gang Lu & Jiashu Cao

  2. Key Laboratory of Horticultural Plant Growth, Development and Quality Improvement, Ministry of Agriculture, Hangzhou, 310029, People’s Republic of China

    Aihong Zhang, Lin Qiu, Li Huang, Xiaolin Yu, Gang Lu & Jiashu Cao

Authors
  1. Aihong Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Lin Qiu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Li Huang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Xiaolin Yu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Gang Lu
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Jiashu Cao
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jiashu Cao.

Electronic Supplementary Materials

Below is the link to the electronic supplementary material.

Online Resource 1

(DOC 66.5 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Zhang, A., Qiu, L., Huang, L. et al. Isolation and Characterization of an Anther-Specific Polygalacturonase Gene, BcMF16, in Brassica campestris ssp. chinensis . Plant Mol Biol Rep 30, 330–338 (2012). https://doi.org/10.1007/s11105-011-0341-2

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11105-011-0341-2

Keywords

Search

Navigation