Poster: Cell Cycle & Cytokinesis
Abs #
1087: Functional analysis of cyclin D3 from Nicotiana tabacum
In eukaryotes, progression of the cell division requires the coordinated activation of cyclin-dependent kinases (CDKs). The retinoblastoma (Rb) protein operates as a negative regulator of cell proliferation in mammalian cells. It binds to the E2F family of transcription factors, and the resulting Rb-E2F complex blocks the transcription of E2F-regulated genes. In response to growth-promoting signals during G1 phase, cyclin D is synthesized to form active complexes with CDK4/6 that phosphorylate Rb, thereby alleviating its repressor function on E2F-controlled gene transcription, and stimulating entry into S phase. Therefore, the Rb pathway plays a pivotal role to control the passage through G1 to S phase. The existence of the key components in plants suggests that the Rb pathway acts a similar role in regulating plant cell cycle. We have isolated two tobacco cyclin (CycD3) genes (NtcycD3-2a, NtcycD3-2b) whose transcripts accumulated in G2-M phase. GST pull-down experiments reveal that both of tobacco CycD3 can bind to CDKA as well as CDKB. By contrast, Arabidopsis CycD3;1 can bind to CDKA but not to CDKB. CDKB represents a plant-specific CDKs involved in the control of G2/M progression, suggesting that tobacco CycD3 may function as a regulatory partner of CDKB in G2/M progression. Using a baculovirus expression system, both CycD3/CDKA and CycD3/CDKB complexes exhibit kinase activity against histone H1 in vitro. In addition, we have prepared the immunoprecipitates from the synchronized tobacco BY-2 cells and these were used for kinase assay on histone H1 and tobacco Rb. This showed that kinase activity of CycD3/CDK complexes exhibited two peaks in late G1/S phase and in G2 phase, suggesting that tobacco CycD3 potentially regulates both G1/S and G2/M progression.
