Proteomic Analysis Reveals a Novel Mutator S (MutS) Partner Involved in Mismatch Repair Pathway
Proteomic Analysis Reveals a Novel Mutator S (MutS) Partner Involved in Mismatch Repair Pathway
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DOI：
10.1074/mcp.m115.056093
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发表时间：
2016-04-01
影响因子：
7
通讯作者：
Chen, Junjie
中科院分区：
文献类型：
--
作者：
Chen, Zhen;Tran, Mykim;Chen, Junjie
错配修复(MMR)家族是一组高度保守的蛋白质，在纠正DNA复制过程中产生的碱基和插入-缺失错配方面发挥功能。这一过程的中断会导致特有的微卫星不稳定(MSI)、修复缺陷和癌症易感性。然而，很大一部分MSI阳性的癌症在正常水平表达MMR基因，并且在已知的MMR基因中没有检测到突变，这表明可能存在其他因素和/或机制来解释患者的这些MSI表型。为了系统地研究MMR途径，我们进行了蛋白质组学分析，并利用串联亲和纯化与质谱联用(TAP-MS)方法鉴定了MMR相关蛋白复合体。质谱学数据已存储到ProteomeXchange中，标识符为PXD003014和DOI 10.6019/PXD003014。我们鉴定了230个高置信度候选相互作用蛋白(HCIP)。随后，我们专注于MMR途径的重要组成部分MSH2，并发现了一个新的MSH2结合伙伴WDHD1。我们进一步证明了WDHD1与MSH2、MSH3或MSH6形成稳定的络合物，即MutS络合物。MSH2/WDHD1的特异性相互作用是由MSH2的第二水平结构域和WDHD1的Ala(1123)位点介导的。此外，我们发现，就像MSH2缺失的细胞一样，WDHD1的缺失也导致了6-硫代鸟嘌呤(6-TG)的抗性，这表明WDHD1可能参与了MMR途径。综上所述，我们的研究揭示了MMR途径中涉及的新成分，该途径提供了可能导致MSI阳性癌症发展的候选基因。
The mismatch repair (MMR) family is a highly conserved group of proteins that function in correcting base-base and insertion-deletion mismatches generated during DNA replication. Disruption of this process results in characteristic microsatellite instability (MSI), repair defects, and susceptibility to cancer. However, a significant fraction of MSI-positive cancers express MMR genes at normal levels and do not carry detectable mutation in known MMR genes, suggesting that additional factors and/or mechanisms may exist to explain these MSI phenotypes in patients. To systematically investigate the MMR pathway, we conducted a proteomic analysis and identified MMR-associated protein complexes using tandem-affinity purification coupled with mass spectrometry (TAP-MS) method. The mass spectrometry data have been deposited to the ProteomeXchange with identifier PXD003014 and DOI 10.6019/PXD003014. We identified 230 high-confidence candidate interaction proteins (HCIPs). We subsequently focused on MSH2, an essential component of the MMR pathway and uncovered a novel MSH2-binding partner, WDHD1. We further demonstrated that WDHD1 forms a stable complex with MSH2 and MSH3 or MSH6, i.e. the MutS complexes. The specific MSH2/WDHD1 interaction is mediated by the second lever domain of MSH2 and Ala(1123) site of WDHD1. Moreover, we showed that, just like MSH2-deficient cells, depletion of WDHD1 also led to 6-thioguanine (6-TG) resistance, indicating that WDHD1 likely contributes to the MMR pathway. Taken together, our study uncovers new components involved in the MMR pathway, which provides candidate genes that may be responsible for the development of MSI-positive cancers.