Scientia Agricultura Sinica ›› 2014, Vol. 47 ›› Issue (20): 3929-3940.doi: 10.3864/j.issn.0578-1752.2014.20.001

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Proteome Analysis of Cytoplasmic Male Sterility and Its Maintaince in JA-CMS Cotton

YANG Peng1,2, HAN Jin-feng1, HUANG Jin-ling3   

  1. 1College of Agronomy, Henan Agricultural University, Zhengzhou 450002
    2 College of Rural Developmen, Shanxi Agricultural University, Taigu 030801, Shanxi
    3College of Agronomy, Shanxi Agricultural University, Taigu 030801, Shanxi
  • Received:2014-04-11 Revised:2014-07-21 Online:2014-10-16 Published:2014-10-16

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Abstract: 【Objective】This study was conducted to analyze and compare proteomes of cotton flower buds at the microspore abortion stage between a cytoplasmic male sterile (CMS) line JA-CMS and its maintainer line JB.【Method】Two-dimensional gel electrophoresis (2-DE) technique was used to separate the protein spots of JA-CMS and JB flower buds at the microspore abortion stage and the gel was stained with Coomassie Blue R-350 solution. Differentially expressed (DE) protein spots were selected with more than 2-fold changes and P values less than 0.05 by PDQuest (version 8.0.1) image software. LC-Chip/ESI-QTOF-MS analysis was carried out to obtain peptide mass fingerprinting of the DE protein spots. The Mascot software was used to search against the NCBInr database for protein annotation. GO (Gene Ontology) and KEGG (Kyoto Encyclopedia of Genes and Genomes) analyses were performed to further explore the biological functions of the DE proteins.【Result】The 2-DE maps of JA-CMS at the sporogenous cells stage (SS) and microsporocyte stage (MS) isolated 1 525 and 1 540 protein spots, respectively, while 1 554 and 1 540 protein spots were detected in JB at the same stages. These protein spots were found within Mr 10-100 kD and pI 3-10. Fifteen DE protein spots between JA-CMS and JB were selected after the quantitative and statistical analysis. Among the 15 proteins, H(+)-transporting ATP synthase, glutathione reductase, ATPase subunit 1 (mitochondrion), ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit, and UDP-D-glucose dehydrogenase were differentially expressed at both the SS and MS stages between JA-CMS and JB. Anx1, S-formylglutathione hydrolase, momilactone A synthase-like, an unnamed protein product with allene oxide cyclase activity, and a conserved hypothetical protein located in mitochondria were down-regulated only at the SS stage in JA-CMS; beta-hydroxyacyl-ACP dehydratase, pyruvate dehydrogenase alpha subunit, and a predicted protein related with pollen development were differentially expressed only at the MS stage between the two materials. Functional category analysis indicated that these DE proteins were mainly involved in processes crucial for microspore and tapetum development. Down- and up-regulation of these proteins may disrupt the coordination of developmental and metabolic processes, resulting in abnormal microspores and defective tapetum which leads to male sterility in JA-CMS ultimately. Quantitative RT-PCR was used to validate Rubisco and H(+)-transporting ATP synthase gene expressions at the transcriptional level, which are consistent with the protein amounts detected dy 2-DE.【Conclusion】CMS phenotype of JA-CMS may be associated with energy metabolism disturbance, abnormal jasmonic acid biosynthesis, accumulation of reactive oxygen species (ROS), and decreased activity of chalcone synthase (CHS). CMS is regulated by a complex signal network, multiple genes of different metabolic pathways may affect pollen fertility.

Key words: cotton, cytoplasmic male sterility (CMS), proteomics, 2-DE

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