A function for cyclin D1 in DNA repair uncovered by protein interactome analyses in human cancers

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Abstract

Cyclin D1 is a component of the core cell cycle machinery1. Abnormally high levels of cyclin D1 are detected in many human cancer types2. To elucidate the molecular functions of cyclin D1 in human cancers, we performed a proteomic screen for cyclin D1 protein partners in several types of human tumours. Analyses of cyclin D1 interactors revealed a network of DNA repair proteins, including RAD51, a recombinase that drives the homologous recombination process3. We found that cyclin D1 directly binds RAD51, and that cyclin D1–RAD51 interaction is induced by radiation. Like RAD51, cyclin D1 is recruited to DNA damage sites in a BRCA2-dependent fashion. Reduction of cyclin D1 levels in human cancer cells impaired recruitment of RAD51 to damaged DNA, impeded the homologous recombination-mediated DNA repair, and increased sensitivity of cells to radiation in vitro and in vivo. This effect was seen in cancer cells lacking the retinoblastoma protein, which do not require D-cyclins for proliferation4,5. These findings reveal an unexpected function of a core cell cycle protein in DNA repair and suggest that targeting cyclin D1 may be beneficial also in retinoblastoma-negative cancers which are currently thought to be unaffected by cyclin D1 inhibition.

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Figure 1: Analyses of cyclin D1 interactors identified in human cancers.
Figure 2: Impaired DNA repair upon reduction of cyclin D1 levels.
Figure 3: Functional interaction between cyclin D1 and RAD51.
Figure 4: Increased radiation-sensitivity of tumours with reduced cyclin D1 levels.

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Acknowledgements

We thank the members of the Sicinski lab for help and advice, N. Li for help with initial experiments, S Panyim and Y. Geng for discussions and reading the manuscript, P. Nakatani for pOZ-FH-N construct, M. Jasin for DR-GFP system, D. Bulavin and E. Appella for anti-phospho-CDC25A antibodies, A. Smogorzewska for anti-FANCI antibody, A. Venkitaraman and M. Lee for GST–BRCA2 fragments, and DFCI Confocal and Light Microscopy Core Facility for assisting with confocal microscopy. This work was supported by R01 CA083688, P01 CA080111 and P01 CA109901 grants from NIH (to P.S.). S.J. is supported by The Thailand Research Fund MRG5280248, W.M. by Foundation for Polish Science, Y.E.W. through the CCCB and the Dana-Farber Strategic Plan Initiative, A.B.C. and T.A.K. by Project Z01 ES065089 (to T.A.K.) from the Division of Intramural Research of the NIH, NIEHS.

Author information

S.J. and P.S. designed the study, analysed the data and wrote the manuscript. S.J. performed most of the experiments with help from collaborators. Y.H. and D.M.L. contributed to DNA repair analyses. W.M. contributed in vitro protein binding analyses. J.E.E. and T.G. performed mass spectrometry; J.E.E. analysed mass spec data with S.P.G. L.B., F.B., A.Z., T.v.H helped with experiments. Y.E.W. M.C. and J.Q. performed computational analyses of interactors. A.B.C. and T.A.K. contributed DNA mismatch repair analyses. B.X. helped with BRCA2 analyses.

Correspondence to Piotr Sicinski.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Figures

This file contains Supplementary Figures 1-28 with legends. (PDF 3085 kb)

Supplementary Information

This file contains Supplementary Methods and additional references. (PDF 972 kb)

Supplementary Table 1

This table contains a list of peptides identified in five human cancer cell lines. (XLS 7218 kb)

Supplementary Table 2

This table contains a list of proteins identified in five human cancer cell lines. (XLS 1191 kb)

Supplementary Table 3

This table contains a list of proteins identified in five human cancer cell lines. (XLS 155 kb)

Supplementary Table 4

This table contains a list of proteins identified in five human cancer cell lines. (XLS 60 kb)

Supplementary Table 5

This table contains a summary of proteomic data for proteins mentioned in the text. (XLS 64 kb)

Supplementary Table 6

This table contains the molecular function biological process analyses of cyclin D1 interacting proteins. (XLS 184 kb)

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