Cholangiocyte microRNAs Regulate Mcl-1 and Cell Death

描述(由申请人提供)： 我的长期职业目标是成为一名独立的研究人员，研究细胞死亡在胆道疾病中的作用。我之前在与细胞死亡相关的细胞和分子生物学方面的研究经验包括我的博士论文，即线粒体突变在年龄相关疾病中的作用。我已经在包括癌症在内的肝胆疾病的细胞死亡领域找到了一位非常成功的导师。该实验室位于罗切斯特梅奥诊所消化系统疾病基础研究中心的有利环境中。这项提案概述了一个可测试的假说，以确定microRNAs在胆管癌细胞中作为细胞凋亡调节器的作用，胆管癌细胞是一种在慢性胆管炎症存在的情况下发生的恶性肿瘤。目前的提案将允许候选人通过提供与细胞死亡相关的肝胆疾病和癌症生物学的高级培训来改变研究方向。短期职业目标包括癌症生物学和Gl相关的翻译研究方面的高级培训。这些直接目标将通过职业发展计划实现，包括研究、沟通技能和指导培训。这项研究计划源于我们之前的观察，即抗凋亡蛋白Mcl-1在人类胆管癌细胞中过度表达，是细胞死亡的关键调节因子。Mcl-1调控的一个潜在机制是通过microRNAs，即充当蛋白质表达序列特异性沉默的短RNAs。我们的初步数据显示，Mcl-1的表达是mir-29b的预测靶点，而mir-29的拮抗作用会增加Mcl-1的蛋白表达。此外，Mcl-1在胆管癌细胞中高表达，而mir-29b表达降低。最后，阻断mir-29可以保护胆管细胞免于细胞死亡。这些广泛和原创性的观察支持了这项拨款的总体假设，即mir-29b的异常表达导致Mcl-1的过度表达，使该肿瘤具有抗凋亡能力。为了解决这一假设，我们建立了先进的细胞培养模型和分子技术来检测和改变microRNAs的表达。这个项目与公共健康相关，因为它研究了胆管癌症如何抵抗细胞死亡，从而抵抗化疗。此外，通过测试一种控制癌细胞死亡的新途径，我们可能会定义一套新的靶点，以改进这种毁灭性疾病的治疗。

DESCRIPTION (provided by applicant): My long-term career goals are to become an independent investigator studying the role of cell death in diseases of the Gl tract. My previous research experience in cell and molecular biology related to cell death included my doctoral dissertation on the role of mitochondrial mutations in age-related disease. I have identified a highly successful mentor in the area of cell death in hepatobiliary diseases, including cancer. The lab is in the conducive environment of the Center for Basic Research in Digestive Diseases, at Mayo Clinic, Rochester. This proposal outlines a testable hypothesis to define the role of microRNAs as regulators of apoptosis in cholangiocarcinoma, a malignant neoplasm that occurs in the presence of chronic inflammation of bile ducts. The current proposal will allow the candidate to change research directions by providing advanced training in hepatobiliary disease and cancer biology as it relates to cell death. Short-term career goals include advanced training in cancer biology and Gl-related translational research. These immediate goals will be achieved by the career development plan, including research, communication skills, and mentoring training. The research plan stems from our previous observations that the antiapoptotic protein Mcl-1 is over-expressed in human cholangiocarcinoma and is a key regulator of cell death. One potential mechanism of Mcl-1 regulation is through microRNAs, short RNAs that act as sequence-specific silencers of protein expression. Our preliminary data shows that Mcl-1 expression is a predicted target of mir-29b, and antagonism of mir-29 increases Mcl-1 protein expression. Further, Mcl-1 is over-expressed in cholangio- carcinoma cells while mir-29b expression is decreased. Finally, blocking mir-29 protects cholangiocytes from cell death. These extensive and original observations support the OVERALL HYPOTHESIS of this grant that dysregulated expression of mir-29b results in over expression of Mcl-1, rendering this cancer apoptosis resistant. To address this hypothesis, we have established advanced cell culture models and molecular techniques to detect and alter expression of microRNAs. This project is relevant to public health as it addresses how bile duct cancers resist cell death, and as such resist chemotherapy. In addition, by testing a new pathway governing cancer cell death, we may define a novel set of targets to improve treatment of this devastating disease.