Genome-Wide Identification and Comparison of Cysteine Proteases in the Pollen Coat and Other Tissues in Maize

Cysteine proteases, belonging to the C1-papain family, play a major role in plant growth and development, senescence, and immunity. There is evidence to suggest that pollen cysteine protease (CP) (ZmCP03) is involved in regulating the anther development and pollen formation in maize. However, there is no report on the genome-wide identification and comparison of in the pollen coat and other tissues in maize. In this study, a total of 38 homologous genes of in maize were identified. Subsequently, protein motifs, conserved domains, gene structures, and duplication patterns of 39 are analyzed to explore their evolutionary relationship and potential functions. The -elements were identified in the upstream sequence of 39 , especially those that are related to regulating growth and development and responding to environmental stresses and hormones. The expression patterns of these genes displayed remarked difference at a tissue or organ level in maize based on the available transcriptome data in the public database. Quantitative reverse transcription PCR (RT-qPCR) analysis showed that was preferably expressed at a high level in maize pollen. Analyses by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot, immunofluorescence and immunogold electron microscopy all validated the cellular localization of ZmCP03 in both the pollen coat and pollen cytoplasm. In addition, 142 genes from Arabidopsis (), rice () and cotton (), together with 39 maize , were retrieved to analyze their evolution by comparing with orthologous genes. The results suggested that ZmCP03 was relatively conservative and stable during evolution. This study may provide a referential evidence on the function of in pollen development and germination in maize.