The online version of this article (doi:10.1186/1476-4598-11-10) contains supplementary material, which is available to authorized users.
The authors declare that they have no competing interests.
JT carried out most of the experiments and participated in the preparation of the manuscript. XW participated in reagent preparation, interpretation of critical data, and manuscript preparation and revision. CH conceived of the study and participated in its design and coordination as well as manuscript preparation. All authors read and approved the final manuscript.
Cisplatin (cis-diamminedichloroplatinum (II), CDDP) and its analogues constitute an important class of anticancer drugs in the treatment of various malignancies; however, its effectiveness is frequently affected by mutations in genes involved in the repair and signaling of cisplatin-induced DNA damage. These observations necessitate a need for a better understanding of the molecular events governing cellular sensitivity to cisplatin.
Here, we show that hMSH5 mediates sensitization to cisplatin-induced DNA damage in human cells. Our study indicates that hMSH5 undergoes cisplatin-elicited protein induction and tyrosine phosphorylation. Silencing of hMSH5 by RNAi or expression of hMSH5 phosphorylation-resistant mutant hMSH5Y742F elevates cisplatin-induced G2 arrest and renders cells susceptible to cisplatin toxicity at clinically relevant doses. In addition, our data show that cisplatin promotes hMSH5 chromatin association and hMSH5 deficiency increases cisplatin-triggered γ-H2AX foci. Consistent with a possible role for hMSH5 in recombinational repair of cisplatin-triggered double-strand breaks (DSBs), the formation of cisplatin-induced hMSH5 nuclear foci is hRad51-dependent.
Collectively, our current study has suggested a role for hMSH5 in the processing of cisplatin-induced DSBs, and silencing of hMSH5 may provide a new means to improve the therapeutic efficacy of cisplatin.
Additional file 1: Figure S1 Representative images of CDDP-induced hMSH5 foci formation at 1, 6, and 24 hrs post treatment. Cells were treated with 10 μM CDDP for 2 hrs and hMSH5 foci formation was analyzed at indicated time points after treatment. Figure S2. Representative images of cisplatin-induced γ-H2AX foci formation in 293T, 293T/f-hMSH5, 293T/f-hMSH5Y742F, and 293T hMSH5 RNAi cells (48 hrs post transfection with pmH1P-Bsd/hMSH5 sh-2). (A) Untreated cells were examined in parallel to establish the basal levels of γ-H2AX signal in these cell lines. (B) Cells were treated with 10 μM cisplatin for 2 hrs followed by γ-H2AX foci analysis at 24 hrs after cisplatin removal. Nuclei are counterstained with DAPI and merged images are provided. Figure S3. Representative images of CDDP-induced γ-H2AX, hRad51, and hMSH5 foci formation in 293T and A549 cells. (A) Analysis of CDDP-triggered γ-H2AX and hMSH5 foci formation. (B) Analysis of CDDP-triggered γ-H2AX and hRad51 foci formation. (C) Analysis of CDDP-triggered hRad51 and hMSH5 foci formation. Consistent with hMSH5 cytoplasmic and nuclear distribution patterns, CDDP-induced hMSH5 foci appear to be present in both cytoplasm and nucleus, whereas CDDP-induced γ-H2AX and hRad51 foci are predominately nuclear. Arrows indicate potential overlaps of two different signals. Figure S4. Effects of hMSH5 Tyr742-to-Phe mutation on its interaction with hMSH4 and c-Abl. (A) Co-IP analysis of the interaction between hMSH5 cp-1 Y742F and hMSH4. The results indicated that hMSH5 cp-1 Y742F could interact with hMSH4 as efficient as hMSH5 cp-1. (B) Co-IP analysis of the interaction between hMSH5Y742F and c-Abl. hMSH5Y742F interacted with c-Abl in the same manner as hMSH5 did. (PDF 6 MB)12943_2011_995_MOESM1_ESM.PDF
Authors’ original file for figure 112943_2011_995_MOESM2_ESM.tiff
Authors’ original file for figure 212943_2011_995_MOESM3_ESM.tiff
Authors’ original file for figure 312943_2011_995_MOESM4_ESM.tiff
Authors’ original file for figure 412943_2011_995_MOESM5_ESM.tiff
Authors’ original file for figure 512943_2011_995_MOESM6_ESM.tiff
Authors’ original file for figure 612943_2011_995_MOESM7_ESM.tiff
Bawa S, Xiao W: A mutation in the MSH5 gene results in alkylation tolerance. Cancer Res. 1997, 57: 2715-2720. PubMed
Nowosielska A, Calmann MA, Zdraveski Z, Essigmann JM, Marinus MG: Spontaneous and cisplatin-induced recombination in Escherichia coli. DNA Repair (Amst). 2004, 3: 719-728. 10.1016/j.dnarep.2004.02.009. CrossRef
Nowosielska A, Marinus MG: Cisplatin induces DNA double-strand break formation in Escherichia coli dam mutants. DNA Repair (Amst). 2005, 4: 773-781. 10.1016/j.dnarep.2005.03.006. CrossRef
Helleday T, Lo J, van Gent DC, Engelward BP: DNA double-strand break repair: from mechanistic understanding to cancer treatment. DNA Repair (Amst). 2007, 6: 923-935. 10.1016/j.dnarep.2007.02.006. CrossRef
Lee TH, Yi W, Griswold MD, Zhu F, Her C: Formation of hMSH4-hMSH5 heterocomplex is a prerequisite for subsequent GPS2 recruitment. DNA Repair (Amst). 2006, 5: 32-42. 10.1016/j.dnarep.2005.07.004. CrossRef
Olive PL: Retention of gammaH2AX foci as an indication of lethal DNA damage. Radiother Oncol. 2011,
Aebi S, Kurdi-Haidar B, Gordon R, Cenni B, Zheng H, Fink D, Christen RD, Boland CR, Koi M, Fishel R, Howell SB: Loss of DNA mismatch repair in acquired resistance to cisplatin. Cancer Res. 1996, 56: 3087-3090. PubMed
Duckett DR, Drummond JT, Murchie AI, Reardon JT, Sancar A, Lilley DM, Modrich P: Human MutSalpha recognizes damaged DNA base pairs containing O6-methylguanine, O4-methylthymine, or the cisplatin-d(GpG) adduct. Proc Natl Acad Sci USA. 1996, 93: 6443-6447. 10.1073/pnas.93.13.6443 PubMedCentralCrossRefPubMed
Fink D, Zheng H, Nebel S, Norris PS, Aebi S, Lin TP, Nehme A, Christen RD, Haas M, MacLeod CL, Howell SB: In vitro and in vivo resistance to cisplatin in cells that have lost DNA mismatch repair. Cancer Res. 1997, 57: 1841-1845. PubMed
Samimi G, Fink D, Varki NM, Husain A, Hoskins WJ, Alberts DS, Howell SB: Analysis of MLH1 and MSH2 expression in ovarian cancer before and after platinum drug-based chemotherapy. Clin Cancer Res. 2000, 6: 1415-1421. PubMed
Neher TM, Bodenmiller D, Fitch RW, Jalal S, Turchi JJ: Novel Irreversible Small Molecule Inhibitors of Replication Protein A Display Single Agent Activity and Synergize with Cisplatin. Mol Cancer Ther. 2011,
Her C, Wu X, Griswold MD, Zhou F: Human MutS homologue MSH4 physically interacts with von Hippel-Lindau tumor suppressor-binding protein 1. Cancer Res. 2003, 63: 865-872. PubMed
- MutS homologue hMSH5: role in cisplatin-induced DNA damage response
Joshua D Tompkins
- BioMed Central
Neu im Fachgebiet Onkologie
e.Med Kampagnen-Visual, Mail Icon II