Submitted to: American Society of Plant Physiologists Meeting
Publication Type: Abstract Only
Publication Acceptance Date: July 1, 1997
Publication Date: N/A
Technical Abstract: Cell Cycle arrest in dormant potato tubers occurs at the G1/G0 phase. In all eukaryotes, progression through the G1/S and G2/M stages of cell division is regulated by a distinct class of proteins known as cyclin- dependent kinases (cdks). These cdks can be identified by using antibody to the conserved PSTAIRE motif of the catalytic subunit or, by affinity purification with a recombinant p13 subunit. Using the PSTAIRE antibody, we have observed a 7-8 fold increase in PSTAIRE-precipitable protein in extracts from non-dormant, growth-arrested meristems vs dormant meristems. When comparing dormant vs actively growing meristems, only a 2-fold increase in PSTAIRE-precipitable protein was observed for actively growing meristems. Based on the current structural models for active cdks, we assume that a regulatory cyclin subunit blocks the PSTAIRE domain, such that, precipitation of an active cdk complex is not complete in actively growing meristems. p13-affinity purified proteins from extracts of non-dormant tuber meristems exhibited no histone H1 kinase activity, whereas, affinity purified proteins from extracts of potato cell cultures did. Additionally, we observed that extracts of potato eyes inhibited the kinase activity of control cdk (p/cyclin B). We have further characterized the inhibitor as a 60-80 kDa glycoprotein based on its gel filtration profile and ability to bind to Con-A. Enzymatic analysis shows that the inhibition is due to a potent ATPase activity. I50 values for the partially purified inhibitor, in standard cdk assays, is approximately 10nM. We will continue working on methods to identify the developmental stages at which active cdks are present in potato tuber meristems.