INDUCTION OF MICROSPORE EMBRYOGENESIS IN BRASSICA-NAPUS L IS ACCOMPANIED BY SPECIFIC CHANGES IN PROTEIN-SYNTHESIS

Culture temperature determines the developmental fate of isolated micr ospores from Brassica napus L.. At 18 degrees C, tricellular pollen de velops, whereas culture at 32 degrees C for 8 h leads to the quantitat ive and synchronous induction of embryogenesis, and ultimately to the formation of embryos. We investigated the changes in protein synthesis that are associated with this 8-h inductive period by using in-situ [ S-35]methionine labeling, followed by two-dimensional (2-D) gel electr ophoretic analysis of the radiolabeled proteins. Qualitative and quant itative computer analyses of 2-D [S-35]methionine protein patterns sho wed six polypeptides specifically labeled under embryogenic culture co nditions. Eighteen polypeptides incorporated [S-35]methionine at a sta tistically significant higher rate under embryogenic culture condition s (32 degrees C) than in the controls (18 degrees C), whereas one prot ein was preferentially labeled under non-embryogenic culture condition s (18 degrees C). These results indicate that only a limited number of proteins detectable in the 2-D gels of microspore extracts are associ ated with the early induction of embryogenesis. The reproducible ident ification of the differentially radiolabeled proteins in the 2-D gels allow the sequencing of representative peptides and the isolation of t he corresponding cDNAs. This may lead to the identification and charac terization of proteins associated with the very first stages of plant embryogenesis.