14-3-3Protein Functional Characterization In Developing Maize Kernels

14-3-3proteins are a group of important regulatory proteins that have hardly beenstudied in the context of maize kernel development, a process of critical significancein both biology and crop breeding. Bioinformatics analysis of maize genomeuncovered twelve14-3-3protein species, among which two14-3-3protein speciesgenes, zmAAU93690.1(AU) and zmGF14-12(GF), exhibited prominent expressionprofiles in maize kernel development. Transcript analysis revealed that more than5000AU-and GF-transcripts were identified, accounting for about1/10of the totalnumber of transcripts of genes correlating to maize kernel development. We coupledthe heterologously expressed AU and GF to an affinity matrix to formchromatographic columns, through which the holoprotein slurry from inbred maizeB73kernels (from6to37day after pollination) was eluted. The species-interactingclient proteins were collected and, subsequently, identified using2-DE in combinationwith LC-MS/MS. A total of77proteins that interacted with AU and/or GF wereidentified and classified into10functional categories. Analyses using Gene Ontology(GO) and Kyoto Encyclopedia of Genes&Genomes (KEGG) revealed that more thanhalf of the client proteins are categorized functionally to be metabolism and proteindestination&storage. While, more than60%(47) of the client proteins interact withboth species, involving in predominantly the fundamental functions such as glycolyticmetabolism, protein destination&storage, and starch synthesis. This suggested thatthe functions of AU and GF are largely overlapping. In addition to these commonfeatures, each species exhibited differential preference toward its client proteins ofunique functionalities: AU tends to bind to proteins involved in disease or defenseprocesses, whilst GF tends to interact with proteins involving in metabolism and cellstructure. In addition, results from both gene transcription and proteomic analysesshowed that AU and GF interacted each other, forming inter-and/or intra-speciescomplexes that participate in various cellular processes during maize kerneldevelopment. Our findings suggest that functional cross-talk likely exista among the 14-3-3proteins in developing maize kernels, and that inter-and/or intra-speciesbindings are likely the primary mechanism underlying the diverse functions of14-3-3proteins. The current results suggested that, for every isoforms, their functionaldifferentiation may be achieved through4possible modes, i.e. clientprotein-determined mode, inter-isoform binding mode, subcellularlocalization-dependent mode and protein post-translational modification mode.