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The APC/C and CBP/p300 cooperate to regulate transcription and cell-cycle progression

Abstract

The anaphase-promoting complex/cyclosome (APC/C) is a multicomponent E3 ubiquitin ligase that, by targeting protein substrates for 26S proteasome-mediated degradation through ubiquitination, coordinates the temporal progression of eukaryotic cells through mitosis and the subsequent G1 phase of the cell cycle1,2,3,4. Other functions of the APC/C are, however, less well defined. Here we show that two APC/C components, APC5 and APC7, interact directly with the coactivators CBP and p300 through protein–protein interaction domains that are evolutionarily conserved in adenovirus E1A5,6,7,8. This interaction stimulates intrinsic CBP/p300 acetyltransferase activity and potentiates CBP/p300-dependent transcription. We also show that APC5 and APC7 suppress E1A-mediated transformation in a CBP/p300-dependent manner, indicating that these components of the APC/C may be targeted during cellular transformation. Furthermore, we establish that CBP is required in APC/C function; specifically, gene ablation of CBP by RNA-mediated interference markedly reduces the E3 ubiquitin ligase activity of the APC/C and the progression of cells through mitosis. Taken together, our results define discrete roles for the APC/C–CBP/p300 complexes in growth regulation.

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Figure 1: Binding of APC5, APC7 and E1A to CBP/p300 through conserved domains.
Figure 2: Role of the APC/C in CBP/p300-regulated transcription.
Figure 3: Suppression of E1A-mediated transformation by APC5 and APC7.
Figure 4: Role of CBP in mitosis.

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Acknowledgements

We thank D. Brockmann, M. J. Campbell, R. T. Hay, D. P. Lane, M. W. Kirschner, T. Kouzarides, W. L. Kraus, E. Moran, J. S. Mymryk and J. Pines for reagents; C. J. McCabe for help with PCR analysis; and J.S.M for help with alignments and for critically reading the manuscript. This work was supported by a Cancer Research UK programme grant. P.H.G. is a Cancer Research UK Gibb Fellow. G.S.S. was supported by a Cancer Research UK career development fellowship and an EMBO long-term fellowship. H.Y. was supported by Marie Curie Cancer Care and the Association for International Cancer Research. S.J.E. is an investigator with the Howard Hughes Medical Institute and is supported by a grant from the NIH.

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Supplementary information

Supplementary Notes

This file contains Supplementary Methods, additional references and legends to accompany the Supplementary Figures. (DOC 51 kb)

Supplementary Table 1

This shows the number of A549 cells in different stages of mitosis following knock-down with a non-silencing siRNA, or following knock-down with an siRNA targeted against CBP. (DOC 42 kb)

Supplementary Figure 1

CBP/p300 association with the APC/C is not dependent on cell cycle status. (PDF 259 kb)

Supplementary Figure 2

E1A homology domains present in both APC5 and APC7 bind to the C/H3 domain of CBP. (PDF 103 kb)

Supplementary Figure 3

The effects of knocking-down APC5 and APC7 gene expression on HSF cell cycle status. (PDF 320 kb)

Supplementary Figure 4

The first part of this figure shows silver-stained gels of the His6-p300 and APC/C preparations used. The second part of the figure shows the effects of knocking-down APC5, APC7, Cdc20 and Cdh1 gene expression by RNAi upon CBP and p300 protein levels in HSFs. (PDF 116 kb)

Supplementary Figure 5

The role of the APC/C, and in particular APC/C components APC5 and APC7, in the regulation of CBP/p300-dependent activation of E2F-1/DP-1. (PDF 619 kb)

Supplementary Figure 6

This figure shows by deconvolution immunofluorescent microscopy that APC5 and APC7 co-localize with CBP in interphase cells and that APC5 also co-localizes with acetylated histone H3 (PDF 1343 kb)

Supplementary Figure 7

GST pull-downs experiments showing CBP/p300 interaction with APC5 mutants and demonstrating that both APC5 and APC7 compete with E1A for binding to CBP. (PDF 181 kb)

Supplementary Figure 8

E1A targets APC5 and APC7 during the transformation process by binding to CBP/p300. (PDF 113 kb)

Supplementary Figure 9

This figure shows by deconvolution immunofluorescent microscopy that APC5 co-localizes with CBP during mitosis in A549 cells. (PDF 960 kb)

Supplementary Figure 10

A proposed model for APC/C-dependent regulation of CBP/p300-directed A-T activity and, moreover, a model for CBP/p300-dependent regulation of the APC/C E3 ubiquitin ligase activity. (PDF 672 kb)

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Turnell, A., Stewart, G., Grand, R. et al. The APC/C and CBP/p300 cooperate to regulate transcription and cell-cycle progression. Nature 438, 690–695 (2005). https://doi.org/10.1038/nature04151

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