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Human FEM1B is required for Rad9 recruitment and CHK1 activation in response to replication stress

Oncogene volume 28, pages 1971–1981 (2009)Cite this article

Abstract

Human checkpoint kinase 1 (CHK1) is an essential kinase required to preserve genome stability, and is activated by DNA replication blockage through the ataxia-telangiectasia-mutated-and-Rad3-related (ATR)/ATRIP-signaling pathway. In this report, we show that a novel CHK1-interacting protein, FEM1B (human homologue of the Caenorhabditis elegans sex determination fem1 protein), identified by a yeast two-hybrid screen, is involved in the activation of CHK1 by replication stress. Depletion of FEM1B by small interfering RNA in cancer cells impairs the activation of CHK1 kinase activity and attenuates the induction of CHK1 Ser345 phosphorylation upon replication interference. It is to be noted that, CHK2 Thr68 phosphorylation is not altered by FEM1B downregulation. By fractionation, we further demonstrated that FEM1B is able to associate with chromatin, and such association facilitates chromatin loading of the Rad9 protein. Consistently, ATR activity is poorly maintained in FEM1B knockdown cells; and FEM1B-ablated cells are as sensitive to replication block as CHK1-depleted cells. Our study has uncovered an adaptor protein FEM1B, which acts as a bridge linking CHK1 and Rad9, thus facilitating checkpoint signaling induced by replication stress.

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Acknowledgements

We thank Shiaw-Wei Tyan for his assistance with yeast library screening. We are also grateful to Drs Hsiu-Ming Shih for the human testis cDNA library; Victor Yu for pXJ-HA-F1Aα; Carol Prives for GST-Cdc25C200−256; Howard Lieberman for Rad9 cDNA; Randal Tibbetts and Robert Abraham for Flag-ATR. This study was supported by funds from Academia Sinica, Taiwan to S-Y Shieh.

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  1. Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan

    T-P Sun

  2. Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan

    T-P Sun & S-Y Shieh

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  1. T-P Sun
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Correspondence to S-Y Shieh.

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Sun, TP., Shieh, SY. Human FEM1B is required for Rad9 recruitment and CHK1 activation in response to replication stress. Oncogene 28, 1971–1981 (2009). https://doi.org/10.1038/onc.2009.58

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