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F-box protein FBXO31 mediates cyclin D1 degradation to induce G1 arrest after DNA damage

Nature volume 459, pages 722–725 (2009)Cite this article

Abstract

In response to DNA damage, eukaryotic cells initiate a complex signalling pathway, termed the DNA damage response (DDR), which coordinates cell cycle arrest with DNA repair. Studies have shown that oncogene-induced senescence, which provides a barrier to tumour development, involves activation of the DDR1,2,3. Using a genome-wide RNA interference (RNAi) screen, we have identified 17 factors required for oncogenic BRAF to induce senescence in primary fibroblasts and melanocytes4. One of these factors is an F-box protein, FBXO31, a candidate tumour suppressor encoded in 16q24.3, a region in which there is loss of heterozygosity in breast, ovarian, hepatocellular and prostate cancers5,6,7,8,9. Here we study the cellular role of FBXO31, identify its target substrate and determine the basis for its growth inhibitory activity. We show that ectopic expression of FBXO31 acts through a proteasome-directed pathway to mediate the degradation of cyclin D1, an important regulator of progression from G1 to S phase, resulting in arrest in G1. Cyclin D1 degradation results from a direct interaction with FBXO31 and is dependent on the F-box motif of FBXO31 and phosphorylation of cyclin D1 at Thr 286, which is known to be required for cyclin D1 proteolysis. The involvement of the DDR in oncogene-induced senescence prompted us to investigate the role of FBXO31 in DNA repair. We find that DNA damage induced by γ-irradiation results in increased FBXO31 levels, which requires phosphorylation of FBXO31 by the DDR-initiating kinase ATM. RNAi-mediated knockdown of FBXO31 prevents cells from undergoing efficient arrest in G1 after γ-irradiation and markedly increases sensitivity to DNA damage. Finally, we show that a variety of DNA damaging agents all result in a large increase in FBXO31 levels, indicating that induction of FBXO31 is a general response to genotoxic stress. Our results reveal FBXO31 as a regulator of the G1/S transition that is specifically required for DNA damage-induced growth arrest.

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Figure 1: Ectopic expression of FBXO31 induces G1 arrest and selective degradation of cyclin D1.
Figure 2: FBXO31-mediated cyclin D1 degradation occurs through the proteasomal pathway.
Figure 3: FBXO31 interacts with and directs ubiquitylation of cyclin D1.
Figure 4: Cell cycle arrest after DNA damage requires ATM-mediated induction of FBXO31.

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Acknowledgements

We thank S. Cook, O. Tetsu, M. Kastan and R. Kumar for reagents; M. Pagano for reagents and advice; L. Chamberlain for technical assistance; and S. Evans for editorial assistance. N.W. is supported in part by a fellowship from the Our Danny Cancer Fund. M.R.G. is an investigator of the Howard Hughes Medical Institute.

Author Contributions M.K.S. and N.W. contributed equally to this work. M.K.S., N.W. and M.R.G. conceived and designed the experiments. M.K.S. and N.W. performed the experiments and, together with M.R.G., analysed the data and wrote the paper.

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  1. Manas K. Santra and Narendra Wajapeyee: These authors contributed equally to this work.

Authors and Affiliations

  1. Howard Hughes Medical Institute, Programs in Gene Function and Expression and Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts 01605, USA ,

    Manas K. Santra, Narendra Wajapeyee & Michael R. Green

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  1. Manas K. Santra
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Correspondence to Michael R. Green.

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Santra, M., Wajapeyee, N. & Green, M. F-box protein FBXO31 mediates cyclin D1 degradation to induce G1 arrest after DNA damage. Nature 459, 722–725 (2009). https://doi.org/10.1038/nature08011

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