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Lamin misexpression upregulates three distinct ubiquitin ligase systems that degrade ATR kinase in HeLa cells

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Abstract

Lamins are the major structural components of the nucleus and mutations in the human lamin A gene cause a number of genetic diseases collectively termed laminopathies. At the cellular level, lamin A mutations cause aberrant nuclear morphology and defects in nuclear functions such as the response to DNA damage. We have investigated the mechanism of depletion of a key damage sensor, ATR (Ataxia-telangiectasia-mutated-and-Rad3-related) kinase, in HeLa cells expressing lamin A mutants or lamin A shRNA. The degradation of ATR kinase in these cells was through the proteasomal pathway as it was reversed by the proteasomal inhibitor MG132. Expression of lamin A mutants or shRNA led to transcriptional activation of three ubiquitin ligase components, namely, RNF123 (ring finger protein 123), HECW2 (HECT domain ligase W2) and the F-box protein FBXW10. Ectopic expression of RNF123, HECW2 or FBXW10 directly resulted in proteasomal degradation of ATR kinase and the ring domain of RNF123 was required for this degradation. However, these ligases did not alter the stability of DNA-dependent protein kinase, which is not depleted upon lamin misexpression. Although degradation of ATR kinase was reversed by MG132, it was not affected by the nuclear export inhibitor, leptomycin B, suggesting that ATR kinase is degraded within the nucleus. Our findings indicate that lamin misexpression can lead to deleterious effects on the stability of the key DNA damage sensor, ATR kinase by upregulation of specific components of the ubiquitination pathway.

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Acknowledgments

The authors would like to thank Bo Cen and Akira Nakagawara for generously providing reagents. The authors are grateful to Nandini Rangaraj for help with confocal microscopy. V. K. P. is a recipient of the J. C. Bose national fellowship from the Department of Science and Technology, India. P. C. and K. S. were supported by predoctoral fellowships from the Council of Scientific and Industrial Research and Department of Biotechnology, respectively.

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  1. Centre for Cellular and Molecular Biology (CSIR), Uppal Road, Hyderabad, 500007, India

    Bhattiprolu Muralikrishna, Pankaj Chaturvedi, Kirti Sinha & Veena K. Parnaik

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  1. Bhattiprolu Muralikrishna
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  2. Pankaj Chaturvedi
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Correspondence to Veena K. Parnaik.

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Muralikrishna, B., Chaturvedi, P., Sinha, K. et al. Lamin misexpression upregulates three distinct ubiquitin ligase systems that degrade ATR kinase in HeLa cells. Mol Cell Biochem 365, 323–332 (2012). https://doi.org/10.1007/s11010-012-1272-4

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