Cell cycle regulation of cyclin-dependent kinases in tobacco cultivar bright yellow-2 cells

Plants possess two major classes of cyclin-dependent kinases (CDK) with cyc lin-binding motifs PSTAIRE (CDK-a) and PPTA/TLRE (CDK-b). Tobacco (Nicotian a tabacum L. cv Bright Yellow-2) cells are the most highly synchronizable p lant culture, but no detailed analysis of CDK activities has been reported in this system. Here we describe isolation of new PPTALRE CDKs (Nicta;CdkB1 ) from Bright Yellow-2 cells and present detailed analysis of the mRNA, pro tein and kinase activity levels of CdkB1, and the PSTAIRE CDKA during the g rowth and cell cycles. CdkA and CdkB1 transcripts are more abundant in expo nential than in stationary phase cells, but the two genes show strikingly d ifferent regulation during the cell cycle. CdkA mRNA and protein accumulate during G1 in cells re-entering the cell cycle, and immunoprecipitated hist one H1 kinase activity increases at the G1/S boundary. Aphidicolin synchron ized cells show the highest CDKA-associated histone H1 kinase activity duri ng S-G2 phases, although CdkA mRNA and protein levels are not significantly regulated. In contrast, CdkB1 transcripts are present at very low levels u ntil S phase and CDKB1 protein and kinase activity is almost undetectable i n G1. CdkB1 mRNA accumulates through S until M phase and its associated kin ase activity peaks at the G2/M boundary, confirming that transcription of P PTALRE CDKs is cell cycle regulated. We suggest that CDKA kinase activity l ikely plays roles at the G1/S phase boundary, during S phase, and at the G2 /M phase transition, and that CDKB1 kinase activity is present only at G2/M .