Dissecting regulatory pathways of G1/S control in Arabidopsis: common and distinct targets of CYCD3;1, E2Fa and E2Fc

Activation of E2F transcription factors at the G1-to-S phase boundary, with the resultant expression of genes needed for DNA synthesis and S-phase, is due to phosphorylation of the retinoblastoma-related (RBR) protein by cyclin D-dependent kinase (CYCD-CDK), particularly CYCD3-CDKA. Arabidopsis has three canonical E2F genes, of which E2Fa and E2Fb are proposed to encode transcriptional activators and E2Fc a repressor. Previous studies have identified genes regulated in response to high-level constitutive expression of E2Fa and of CYCD3;1, but such plants display significant phenotypic abnormalities. We have sought to identify targets that show responses to lower level induced changes in abundance of these cell cycle regulators. Expression of E2Fa, E2Fc or CYCD3;1 was induced using dexamethasone and the effects analysed using microarrays in a time course allowing short and longer term effects to be observed. Overlap between CYCD3;1 and E2Fa modulated genes substantiates their action in a common pathway with a key role in controlling the G1/S transition, with additional targets for CYCD3;1 in chromatin modification and for E2Fa in cell wall biogenesis and development. E2Fc induction led primarily to gene downregulation, but did not antagonise E2Fa action and hence E2Fc appears to function outside the CYCD3-RBR pathway, does not have a direct effect on cell cycle genes, and promoter analysis suggests a distinct binding site preference.