Plant Molecular Biology

, 71:345 | Cite as

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

  • Sarah M. de Jager
  • Simon Scofield
  • Rachael P. Huntley
  • Alastair S. Robinson
  • Bart G. W. den Boer
  • James A. H. Murray
Article

Abstract

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.

Keywords

Arabidopsis thaliana Cell cycle CYCD E2F Microarray 

Notes

Acknowledgments

This work was funded by the BBSRC, Cambridge Commonwealth Trust and the European Research Area in Plant Genomics (ERA-PG) project “Plant Stem Cell Network”.

Supplementary material

11103_2009_9527_MOESM1_ESM.xls (2.4 mb)
Table S1: CYCD3;1_All. Pairwise comparison data of microarray analysis of dex induction of CYCD3;1 for 6 h, 24 h and 14 days compared to the empty vector control line treated with dex for the same time periods. Column headings and their description are provided in a separate text document Table S1_columnheadings (XLS 2,473 kb)
11103_2009_9527_MOESM2_ESM.doc (48 kb)
Supplementary material 14 (DOC 47 kb)
11103_2009_9527_MOESM3_ESM.xls (2.8 mb)
Table S2: E2Fa/DPa_All. Pairwise comparison data of microarray analysis of dex induction of E2Fa/DPa for 6 h, 24 h and 14 days compared to the empty vector control line treated with dex for the same time periods. Column headings and their description are provided in a separate text document Table S2 columnheadings (XLS 2,888 kb)
11103_2009_9527_MOESM4_ESM.doc (50 kb)
Supplementary material 15 (DOC 50 kb)
11103_2009_9527_MOESM5_ESM.xls (1.6 mb)
Table S3: E2Fc/DPa_All. Pairwise comparison data of microarray analysis of dex induction of E2Fc/DPa for 6 h, 24 h and 14 days compared to the empty vector control line treated with dex for the same time periods. Column headings and their description are provided in a separate text document Table S3 columnheadings(XLS 1,629 kb)
11103_2009_9527_MOESM6_ESM.doc (50 kb)
Supplementary material 16 (DOC 50 kb)
11103_2009_9527_MOESM7_ESM.xls (48 kb)
Table S4: CYCD3;1 overlaps. Genes robustly modulated after dex induction of CYCD3;1 for 6 h, 24 h and 14 days which show overlap between any of the three time points are listed. Whether the gene is called increased (I) or decreased (D) is given for each of the three time points, together with the MIPS code and gene description (XLS 48 kb)
11103_2009_9527_MOESM8_ESM.xls (58 kb)
Table S5: E2Fa/DPa overlaps. Genes robustly modulated after dex induction of E2Fa/DPa for 6 h, 24 h and 14 days which show overlap between any of the three time points are listed. Whether the gene is called increased (I) or decreased (D) is given for each of the three time points, together with the MIPS code and gene description (XLS 57 kb)
11103_2009_9527_MOESM9_ESM.xls (38 kb)
Table S6: E2Fc/DPa overlaps. Genes robustly modulated after dex induction of E2Fc/DPa for 6 h,24 h and 14 d which show overlap between any of the three time points are listed. Whether the gene is called increased (I) or decreased (D) is given for each of the three time points, together with the MIPS code and gene description (XLS 38 kb)
11103_2009_9527_MOESM10_ESM.xls (26 kb)
Table S7: 6 h overlaps. Genes robustly modulated after dex induction of CYCD3;1, E2Fa/DPa and E2Fc/DPa for 6 h which show overlap between any of the three transgenes are listed. Whether the gene is called increased (I) or decreased (D) is given for each of the three transgenes, together with the MIPS code and gene description (XLS 26 kb)
11103_2009_9527_MOESM11_ESM.xls (171 kb)
Table S8: 24 h overlaps. Genes robustly modulated after dex induction of CYCD3;1, E2Fa/DPa and E2Fc/DPa for 24 h which show overlap between any of the three transgenes are listed. Whether the gene is called increased (I) or decreased (D) is given for each of the three transgenes, together with the MIPS code and gene description (XLS 171 kb)
11103_2009_9527_MOESM12_ESM.xls (92 kb)
Table S9: 14 d Overlaps. Genes robustly modulated after dex induction of CYCD3;1, E2Fa/DPa and E2Fc/DPa for 14 d which show overlap between any of the three transgenes are listed. Whether the gene is called increased (I) or decreased (D) is given for each of the three transgenes, together with the MIPS code and gene description (XLS 91 kb)
11103_2009_9527_MOESM13_ESM.xls (147 kb)
Table S10: NNNSSCGSSNNN. Robustly modulated genes (reduced dataset) after dex induction of CYCD3;1, E2Fa/DPa and E2Fc/DPa transgenes for 6 h, 24 h and 14 d which have the palindromic E2F-site NNNSSCGSSNNN (where N = A/T/C/G and S = C/G) in the first 500 bp of promoter sequence are listed. The number of sites per promoter, the start and end of the sequence, and the sequence are given (XLS 147 kb)
11103_2009_9527_MOESM14_ESM.xls (316 kb)
Table S11: NNNSSCGS. Robustly modulated genes (reduced dataset) after dex induction of CYCD3;1, E2Fa/DPa and E2Fc/DPa transgenes for 6 h, 24 h and 14 d which have the minimal E2F-site NNNSSCGS (where N = A/T/C/G and S = C/G) in the first 500 bp of promoter sequence are listed. The number of sites per promoter, the start and end of the sequence, and the sequence are given. The distribution of the number of gene promoters with core E2F sites and the number of sites analysed as related to the number of sites per promoter is shown (XLS 316 kb)
11103_2009_9527_MOESM15_ESM.xls (35 kb)
Table S12: All_E2F/DPa_arrays. Comparison of genes robustly modulated after dex induction of E2Fa/DPa for 6 h, 24 h and 14 d and as well as in previously published cDNA and micro-array data (Vlieghe et al. 2003; Vandepoele et al. 2005, Naouar et al. 2009). Genes called increased (I) in at least one of the three timepoints as well as reported increased in at least one of the previous published datasets are listed. The presence of E2F sites of the sequences TTTSSCGS, NNNSSCGS and NNNSSCGSSNNN (where N = A/T/C/G and S = C/G) in the 500 bp upstream sequence is indicated (XLS 35 kb)
11103_2009_9527_MOESM16_ESM.xls (2.8 mb)
Table S13: Modulated Candidates. Genes not robustly modulated after dex induction of CYCD3;1, E2Fa/DPa and E2Fc/DPa for 6 h, 24 h and 14 d, but that are “modulation candidates” since they meet more relaxed criteria as defined in section "Materials and methods" are listed. Column headings and their description are provided in a separate text document Table S13 columnheadings (XLS 2,849 kb)
11103_2009_9527_MOESM17_ESM.doc (75 kb)
Supplementary material 17 (DOC 75 kb)

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Copyright information

© Springer Science+Business Media B.V. 2009

Authors and Affiliations

  • Sarah M. de Jager
    • 1
  • Simon Scofield
    • 1
    • 4
  • Rachael P. Huntley
    • 2
  • Alastair S. Robinson
    • 1
  • Bart G. W. den Boer
    • 3
  • James A. H. Murray
    • 1
    • 4
  1. 1.Department of Chemical Engineering and BiotechnologyUniversity of CambridgeCambridgeUK
  2. 2.EMBL-European Bioinformatics InstituteHinxtonCambridgeUK
  3. 3.Bayer BioScience N.V.GhentBelgium
  4. 4.School of BiosciencesCardiff UniversityCardiffWales, UK

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