Effect of the R406H Substitution on the Normal Function of CHEK2


CHEK2 (Checkpoint kinase homolog 2) encodes a protein involved in pathways that arrest the cell cycle in response to genomic stress such as DNA damage or replication blocks. Carriers of some of the CHEK2 mutations are at an increased risk of breast cancer. A mutation in the kinase domain of the CHEK2 gene resulting in the R406H substitution has been reported. However, it is currently unknown whether the substitution alters the function of CHEK2 in vitro. We evaluated the effect of the R406H substitution on the normal function of CHEK2 using a yeast complementation assay. The yeast cells lacking Rad53, the yeast homologue of human CHEK2 were transformed with the wild type as well as plasmids carrying mutations resulting in the R406H, 1100delC, and S428F variants. Yeast cells carrying the R406H variant grew at a rate similar to those carrying the wild type CHEK2, whereas the yeast carrying the S428F and 1100delC mutants grew at a slower rate. These results suggest that, unlike the well-known pathogenic alleles such as 1100delC and S428F, the R406H substitution does not abrogate the function of CHEK2. Therefore, this variant is probably not important in development of breast cancer in women.

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Yilmaz, A. , Davis, M. and Zhao, W. (2014) Effect of the R406H Substitution on the Normal Function of CHEK2. Advances in Bioscience and Biotechnology, 5, 386-392. doi: 10.4236/abb.2014.54046.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] Jemal, A., Siegel, R., Ward, E., Murray, T., Xu, J., Smigal, C. and Thun, M.J. (2006) Cancer Statistics. CA: A Cancer Journal for Clinicians, 56, 106-130. http://dx.doi.org/10.3322/canjclin.56.2.106
[2] Garcia, M.J. and Benitez, J. (2008) The Fanconi Anaemia/BRCA Pathway and Cancer Susceptibility. Searching for New Therapeutic Targets. Clinical and Translational Oncology, 10, 78-84.
[3] Laitman, Y., Kaufman, B., Lahad, E.L., Papa, M.Z. and Friedman, E. (2007) Germline CHEK2 Mutations in Jewish Ashkenazi Women at High Risk for Breast Cancer. Israel Medical Association Journal, 9, 791-796.
[4] Bell, D.W., Varley, J.M., Szydlo, T.E., Kang, D.H., Wahrer, D.C.R., Shannon, K.E., Lubratovich, M., Verseli, S.J., Isselbacher, K.J., Fraumeni, J.F., Birch, J.M., Garber, J.E. and Haber, D.A. (1999) Heterozygous Germ Line hCHK2 Mutations in Li-Fraumeni Syndrome. Science, 286, 2528-2531. http://dx.doi.org/10.1126/science.286.5449.2528
[5] Liang, F. and Wang, Y. (2007) DNA Damage Checkpoints Inhibit Mitotic Exit by Two Different Mechanisms. Molecular and Cellular Biology, 27, 5067-5078. http://dx.doi.org/10.1128/MCB.00095-07
[6] Weischer, M., Bojesen, S.E., Ellervik, C., Tybjaerg-Hansen, A. and Nordestgaard, B.G. (2008) CHEK2*1100delC Genotyping for Clinical Assessment of Breast Cancer Risk: Meta-Analyses of 26,000 Patient Cases and 27,000 Controls. Journal of Clinical Oncology, 26, 542-548. http://dx.doi.org/10.1200/JCO.2007.12.5922
[7] Williams, L.H., Choong, D., Johnson, S.A. and Campbell, I.G. (2006) Genetic and Epigenetic Analysis of CHEK2 in Sporadic Breast, Colon, and Ovarian Cancers. Clinical Cancer Research, 12, 6967-6972.
[8] Cybulski, C., Gliniewicz, B., Sikorski, A., Kñadny, J., Huzarski, T., Gronwald, J., Byrski, T., Debniak, T., Gorski, B., Jakubowska, A., Wokolorczyk, D., Narod, S.A. and Lubiñski, J. (2007) Epistatic Relationship between the Cancer Susceptibility Genes CHEK2 and p27. Cancer Epidemiology, Biomarkers & Prevention, 16, 572-576.
[9] Wasielewski, M., Vasen, H., Wijnen, J., Hooning, M., Dooijes, D., Tops, C., Klijn, J.G., Meijers-Heijboer, H. and Schutte, M. (2008) CHEK2 1100delC Is a Susceptibility Allele for HNPCC-Related Colorectal Cancer. Clinical Cancer Research, 14, 4989-4994. http://dx.doi.org/10.1158/1078-0432.CCR-08-0389
[10] Zlowocka, E., Cybulski, C., Górski, B., Debniak, T., Slojewski, M., Wokolorczyk, D., Serrano-Fernández, P., Matyjasik, J., van de Wetering, T., Sikorski, A., Scott, R.J. and Lubiński, J. (2008) Germline Mutations in the CHEK2 Kinase Gene Are Associated with an Increased Risk of Bladder Cancer. International Journal of Cancer, 122, 583-586.
[11] Soumittra, N., Meenakumari, B., Parija, T., Sridevi, V., Nancy, K.N., Swaminathan, R., Rajalekshmy, K.R., Majhi, U. and Rajkumar, T. (2009) Molecular Genetics Analysis of Hereditary Breast and Ovarian Cancer Patients in India. Hereditary Cancer in Clinical Practice, 7, 13. http://dx.doi.org/10.1186/1897-4287-7-13
[12] Novak, D.J., Chen, L.Q., Ghadirian, P., Hamel, N., Zhang, P., Rossiny, V., Cardinal, G., Robidoux, A., Tonin, P.N., Rousseau, F., Narod, S.A. and Foulkes, W.D. (2008) Identification of a Novel CHEK2 Variant and Assessment of Its Contribution to the Risk of Breast Cancer in French Canadian Women. BMC Cancer, 8, 239.
[13] Chen, W., Yurong, S. and Liansheng, N. (2008) Breast Cancer Low-Penetrance Allele 1100delC in the CHEK2 Gene: Not Present in the Chinese Familial Breast Cancer Population. Advances in Therapy, 25, 496-501.
[14] Zhang, S., Phelan, C.M., Zhang, P., Rousseau, F., Ghadirian, P., Robidoux, A., Foulkes, W., Hamel, N., McCready, D., Trudeau, M., Lynch, H., Horsman, D., De Matsuda, M.L., Aziz, Z., Gomes, M., Costa, M.M., Liede, A., Poll, A., Sun, P. and Narod, S.A. (2008) Frequency of the CHEK2 1100delC Mutation among Women with Breast Cancer: An International Study. Cancer Research, 68, 2154-2157. http://dx.doi.org/10.1158/0008-5472.CAN-07-5187
[15] Zhao, X. and Rothstein, R. (2002) The Dun1 Checkpoint Kinase Phosphorylates and Regulates the Ribonucleotide Reductase Inhibitor Sml1. Proceedings of the National Academy of Sciences of the USA, 99, 3746-3751.
[16] Cordon-Preciado, V., Ufano, S. and Bueno, A. (2006) Limiting Amounts of Budding Yeast Rad53 S-Phase Checkpoint Activity Results in Increased Resistance to DNA Alkylating Damage. Nucleic Acid Research, 34, 5852-5862.
[17] Matsuoka, S., Huang, M. and Elledge, S.J. (1998) Linkage of ATM to Cell Cycle Regulation by the Chk2 Protein Kinase. Science, 282, 1893-1897. http://dx.doi.org/10.1126/science.282.5395.1893
[18] Cline, J., Braman, J.C. and Hogrefe, H.H. (1996) PCR Fidelity of pfu DNA Polymerase and Other Thermostable DNA Polymerases. Nucleic Acids Research, 24, 3546-3551. http://dx.doi.org/10.1093/nar/24.18.3546
[19] Elble, R. (1992) A Simple and Efficient Procedure for Transformation of Yeasts. Biotechniques, 13, 18-20.
[20] Gietz, R.D. and Woods, R.A. (2006) Yeast Transformation by the LiAc/SS Carrier DNA/PEG Method. Methods in Molecular Biology, 313, 107-120.
[21] Tischkowitz, M.D., Yilmaz, A., Chen, L.Q., Karyadi, D.M., Novak, D., Kirchhoff, T., Hamel, N., Tavtigian, S.V., Kolb, S., Bismar, T.A., Aloyz, R., Nelson, P.S., Hood, L., Narod, S.A., White, K.A., Ostrander, E.A., Isaacs, W.B., Offit, K., Cooney, K.A., Stanford, J.L. and Foulkes, W.D. (2008) Identification and Characterization of Novel SNPs in CHEK2 in Ashkenazi-Jewish Men with Prostate Cancer. Cancer Letters, 270, 173-180.
[22] Shaag, A., Walsh, T., Renbaum, P., Kirchhoff, T., Nafa, K., Shiovitz, S., Mandell, J.B., Welcsh, P., Lee, M.K., Ellis, N., Offit, K., Levy-Lahad, E. and King, M.C. (2005) Functional and Genomic Approaches Reveal an Ancient CHEK2 Allele Associated with Breast Cancer in the Ashkenazi Jewish Population. Human Molecular Genetics, 14, 555-563.
[23] Pegg, A.E. (1984) Methylation of the O6 Position of Guanine in DNA Is the Most Likely Initiating Event in Carcinogenesis by Methylating Agents. Cancer Investigation, 2, 223-231. http://dx.doi.org/10.3109/07357908409104376
[24] Jansson, M., Durant, S.T., Cho, E.C., Sheahan, S., Edelmann, M., Kessler, B. and La Thangue, N.B. (2008) Arginine Methylation Regulates the p53 Response. Nature Cell Biology, 10, 1431-1439. http://dx.doi.org/10.1038/ncb1802
[25] Betts, M.J. and Russell, R.B. (2003) Amino Acid Properties and Consequences of Subsitutions. In: Gray, I.C., Ed., Bioinformatics for Geneticists Barnes MR, Wiley. http://dx.doi.org/10.1002/0470867302.ch14
[26] Nurse, P. (2000) A Long Twentieth Century of the Cell Cycle and Beyond. Cell, 100, 71-78.
[27] Sanchez, Y., Desany, B.A., Jones, W.J., Liu, Q., Wang, B. and Elledge, S.J. (1996) Regulation of Rad53 by the ATM-Like Kinases MEC1 and TEL1 in Yeast Cell Cycle Checkpoint Pathways. Science, 271, 357-360.
[28] Simon, I., Barnett, J., Hannett, N., Harbison, C.T., Rinaldi, N.J., Volkert, T.L., Wyrick, J.J., Zeitlinger, J., Gifford, D.K., Jaakkola, T.S. and Young, R.A. (2001) Serial Regulation of Transcriptional Regulators in the Yeast Cell Cycle. Cell, 106, 697-708. http://dx.doi.org/10.1016/S0092-8674(01)00494-9
[29] Visintin, C., Tomson, B.N., Rahal, R., Paulson, J., Cohen, M., Taunton, J., Amon, A. and Visintin, R. (2008) APC/CCdh1-Mediated Degradation of the Polo Kinase Cdc5 Promotes the Return of Cdc14 into the Nucleolus. Genes & Development, 22, 79-90. http://dx.doi.org/10.1101/gad.1601308
[30] Listovsky, T., Zor, A., Laronne, A. and Brandeis, M. (2000) Cdk1 Is Essential for Mammalian Cyclosome/APC Regulation. Experimental Cell Research, 255, 184-191. http://dx.doi.org/10.1006/excr.1999.4788

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