Nuclear heterogeneous nuclear ribonucleoprotein D is associated with poor prognosis and interactome analysis reveals its novel binding partners in oral cancer.
Nuclear heterogeneous nuclear ribonucleoprotein D is associated with poor prognosis and interactome analysis reveals its novel binding partners in oral cancer.
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DOI：
10.1186/s12967-015-0637-3
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发表时间：
2015-08-30
影响因子：
7.4
通讯作者：
Ralhan R
中科院分区：
文献类型：
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作者：
Kumar M;Matta A;Masui O;Srivastava G;Kaur J;Thakar A;Shukla NK;RoyChoudhury A;Sharma M;Walfish PG;Michael Siu KW;Chauhan SS;Ralhan R
异质性核糖核蛋白（hnRNP）的转录后调节是癌症发展中的重要调节模式。我们的蛋白质组学分析显示hnRNPD过度表达在口腔异型增生与正常粘膜相比，其在口腔癌的发生作用仍然未知。本研究旨在探讨口腔鳞状细胞癌中hnRNPD相关蛋白网络的表达及其临床意义。免疫沉淀（IP），然后串联质谱法被用来确定口腔癌细胞系中的hnRNPD的结合伴侣。进行抗干扰途径分析（IPA）以解开与hnRNPD相关的蛋白质相互作用网络，并通过co-IP-western印迹确认关键相互作用。采用免疫组化（IHC）分析183例OSCC、44例口腔异型增生和106例正常组织中hnRNPD的表达，并将其与临床病理参数和91个月的随访数据相关联。Kaplan-Meier生存分析和Cox多因素回归分析用于评估hnRNPD在OSCC中的预后意义。我们在口腔癌细胞中鉴定了345个hnRNPD的结合伴侣。IPA揭示了与hnRNPD相关的新型蛋白质-蛋白质相互作用网络，并表明其参与多个细胞过程：DNA修复，复制，染色质重塑，细胞增殖，RNA剪接和稳定性，从而指导口腔癌细胞的命运。hnRNPD与14-3-3 β、hnRNPK和S100 A9的蛋白质-蛋白质相互作用使用co-IP-western印迹法证实。IHC分析显示，与正常粘膜相比，口腔发育不良[p = 0.001，比值比（OR）= 5.1，95% CI = 2.1-11.1）和OSCC（p = 0.001，OR = 8.1，95% CI = 4.5-14.4）中核hnRNPD显著过表达。通过Kaplan-Meier生存率和Cox多变量回归分析，显示核hnRNPD过表达的OSCC患者的无复发生存率显著降低[p = 0.026，风险比= 1.95，95% CI = 1.0-3.5]，并且有可能在淋巴结阴性疾病的OSCC患者中定义高风险亚组。我们的研究结果表明，除了RNA剪接和口腔癌中的稳定性外，hnRNPD在细胞增殖和存活中具有新的功能。核hnRNPD与OSCC患者预后不良的关系及其在口腔癌中的相关蛋白网络，值得进一步研究，旨在探索其作为分子治疗的一个合理的新靶点的潜力。本文的在线版本（doi：10.1186/s12967-015-0637-3）包含补充材料，可供授权用户使用。
Post-transcriptional regulation by heterogeneous ribonucleoproteins (hnRNPs) is an important regulatory paradigm in cancer development. Our proteomic analysis revealed hnRNPD overexpression in oral dysplasia as compared with normal mucosa; its role in oral carcinogenesis remains unknown. Here in we determined the hnRNPD associated protein networks and its clinical significance in oral squamous cell carcinoma (OSCC). Immunoprecipitation (IP) followed by tandem mass spectrometry was used to identify the binding partners of hnRNPD in oral cancer cell lines. Ingenuity pathway analysis (IPA) was carried out to unravel the protein interaction networks associated with hnRNPD and key interactions were confirmed by co-IP-western blotting. hnRNPD expression was analyzed in 183 OSCCs, 44 oral dysplasia and 106 normal tissues using immunohistochemistry (IHC) and correlated with clinico-pathological parameters and follow up data over a period of 91 months. Kaplan–Meier survival and Cox-multivariate-regression analyses were used to evaluate the prognostic significance of hnRNPD in OSCC. We identified 345 binding partners of hnRNPD in oral cancer cells. IPA unraveled novel protein–protein interaction networks associated with hnRNPD and suggested its involvement in multiple cellular processes: DNA repair, replication, chromatin remodeling, cellular proliferation, RNA splicing and stability, thereby directing the fate of oral cancer cells. Protein–protein interactions of hnRNPD with 14-3-3ζ, hnRNPK and S100A9 were confirmed using co-IP-western blotting. IHC analysis showed significant overexpression of nuclear hnRNPD in oral dysplasia [p = 0.001, Odds ratio (OR) = 5.1, 95 % CI = 2.1–11.1) and OSCCs (p = 0.001, OR = 8.1, 95 % CI = 4.5–14.4) in comparison with normal mucosa. OSCC patients showing nuclear hnRNPD overexpression had significantly reduced recurrence free survival [p = 0.026, Hazard ratio = 1.95, 95 % CI = 1.0–3.5] by Kaplan–Meier survival and Cox-multivariate-regression analyses and has potential to define a high-risk subgroup among OSCC patients with nodal negative disease. Our findings suggest novel functions of hnRNPD in cellular proliferation and survival, besides RNA splicing and stability in oral cancer. Association of nuclear hnRNPD with poor prognosis in OSCC patients taken together with its associated protein networks in oral cancer warrant future studies designed to explore its potential as a plausible novel target for molecular therapeutics. The online version of this article (doi:10.1186/s12967-015-0637-3) contains supplementary material, which is available to authorized users.