Determination of molecular pathways regulating LOT1 mediated growth suppressions

描述：(申请人描述)我们先前通过分析正常和恶性转化的大鼠卵巢表面上皮(ROSE)细胞(一种体外卵巢癌模型)的基因表达差异，发现了LOT1(Lost On Transform)基因(GenBank登录号：U72620)。在独立转化的玫瑰细胞系中，与正常的祖细胞相比，该基因的表达经常减少或丢失。基于这些观察，我们认为LOT1基因是参与调控人卵巢癌的潜在候选基因。为了将模型系统中的这一发现转化为人类疾病，我们克隆了人类同源物LOT1(GenBank Accession#72621)。有趣的是，人类基因定位在6号染色体的25号带(6q25)上，这是卵巢癌和许多其他实体肿瘤中杂合性缺失(LOH)的位置。在11个卵巢癌细胞系中，有7个LOT1基因的表达缺失或降低，这与在大鼠卵巢癌细胞系中的发现一致。推导的氨基酸序列表明，LOT1是一个含锌指基序的蛋白质，具有转录因子结构域的特征。功能分析证明它是一种核蛋白，具有转录因子的潜在作用。我们还发现LOT1基因参与了表皮生长因子受体(EGFR)信号通路，是卵巢癌细胞生长的负调控因子。这项建议的目的是通过确定LOT1的下调或表达/功能缺失如何传递抗增殖信号，以及作为肿瘤/生长抑制基因调控其他抑制恶性表型的基因，进一步确定LOT1的临床重要性。为了阐明这些机制，我们将研究与LOT蛋白的作用相关的上游调节因子和下游事件，并确定这些正常过程在卵巢恶性肿瘤中是如何受到干扰的。希望这些努力将有助于更好地了解卵巢癌的发展和/或进展，并为更有效地诊断、治疗和/或管理这种疾病提供基础。

DESCRIPTION: (Applicant's Description) We previously reported discovery of the LOT1 (Lost On Transformation) gene (GenBank accession #U72620) through analysis of gene expression differences between normal and malignantly transformed rat ovarian surface epithelial (ROSE) cells, an in vitro ovarian cancer model. The gene frequently shows decreased or lost expression in independently transformed ROSE cell lines compared to the normal progenitor cells. Based on these observations, we considered the LOT1 gene a potential candidate gene involved in regulating human ovarian cancer. In an effort to translate this finding in a model system to human disease, we cloned the human homologue, LOT1 (GenBank accession #72621). Interestingly, the human gene maps to chromosome 6 at band 25 (6q25), which is a site for loss of heterozygosity (LOH) in ovarian cancer and many other solid tumors. Expression of the LOT1 gene was lost or decreased in seven out of eleven ovarian cancer cell lines consistent with the findings in the rat ovarian cancer cell lines. The deduced amino acid sequence of LOT1 indicates that it is a zinc-finger motif containing protein with domains characteristic of transcription factors. Functional assays proved that it is a nuclear protein with a potential role as a transcription factor. We also found that the LOT1 gene is involved in the epidermal growth factor receptor (EGFR) signaling pathway and is a negative regulator of the ovarian cancer cell growth. The goal of this proposal is to further ascertain the clinical importance of LOT1 by determining how down-regulation or lost expression/function of LOT1 transduces anti-proliferative signals and behaves as a tumor/growth suppresser gene regulating other genes, which inhibit the malignant phenotype. To elaborate these mechanisms we will investigate the upstream regulators and downstream events associated with the action of the LOT protein and to determine how these normal processes are perturbed in ovarian malignancies. Hopefully, these endeavors will result in a better understanding of the ovarian cancer development and/or progression and provide a basis for more effective diagnosis, treatment, and/or management of the disease.