Proteomic Analysis Of Cytoplasmic Male Sterile Anthers And Functional Analysis Of CaSEP5 Gene In Pepper(Capsicum Annuum L.)

Although cytoplasmic male sterility(CMS)is widely used for developing pepper hybrids,its molecular mechanism remains unclear.In this study,a comparative proteomic analysis was performed between a pepper CMS line(A-line)and its isogenic maintainer line(B-line)via label-free method.Approximately 324 differentially abundant protein species were identified and quantified,among which,47 were up-accumulated,whereas 140 were down-accumulated in the A-line.Additionally,75 and 62 protein species were specifically accumulated in the A-line and B-line,respectively.Protein species involved in pollen exine formation,pyruvate metabolic processes,the tricarboxylic acid cycle,the mitochondrial electron transport chain,and oxidative stress response were observed to be differentially accumulated between A-line and B-line,suggesting their potential roles in the regulation of pepper pollen abortion.Flowers are reproductive organs that determine final fruit or seed yield in flowering plants.SEP-like MADS-box genes participate in floral determination and/or flower organ specification.Nevertheless,their molecular mechanisms in bell pepper have not yet been elucidated.In this study,we identified and cloned five SEP-like genes from pepper flower buds,designated as CaSEPl to CaSEP5(GenBank Accession No.MG745434-38),respectively.Phylogenetic analysis indicated that CaSEP1 belonged to the SEP1/2-subclade,CaSEP2 and CaSEP3 belonged to the SEP4-subclade,CaSEP4 and CaSEP5 belonged to the SEP3-and FBP9/23-subclade,respectively.Repressing the expression of CaSEP-like genes via virus-induced silencing indicated that only CaSEP5-silenced plants showed obvious phenotype,such as elongated and leaf-like sepals,rigid and straight pedicels and reduced fruit size.The expression patterns of other CaSEP-like genes and the B-,C-class genes were altered in CaSEP5-silenced flowers.In another aspect,IAA,ABA,and MeJA levels were significantly reduced in CaSEP5-silenced flower buds,whereas,the IAA and ABA levels in the CaSEP5-silenced pedicels were significantly increased.Expression of CaSEP5 gene had been detected in all whorls of floral organs and was relatively high in sepals and petals.Subcellular localization revealed that CaSEP5 located in the nucleus,which is consistent with the location of a typical transcription factor.Yeast two-hybrid and bimolecular fluorescence complementation results indicated that CaSEP5 may function as a glue to form protein complexes in the regulation of flower development.Based on the results mentioned above,it can be presumed that CaSEP5 plays an important role in pepper flower development by means of a complex phytohormone and genetic network.