Characterization of a Novel Growth Regulator

克隆了一个新的基因，命名为NNGR(新型负生长调节因子)。NNGR编码一种分泌的蛋白质，含有两个锌指状基序，并与酵母杀手毒素同源。NNGR位于染色体4q21.1-22，该区域在食道癌和结直肠癌中经常缺失。与相应的正常组织相比，NNGR在5/6的结肠、1/1的胃和6/7的食道肿瘤中表达缺失或降低。最重要的是，在NNGR不表达的结直肠癌细胞中，NNGR的表达通过诱导细胞凋亡来抑制其生长。因此，NNGR作为NOEL生长抑制因子，在原发胃肠道(GI)癌中表达缺失或降低。这些研究主要针对NNGR的分子和功能特征。具体来说，将利用新鲜的冷冻和石蜡包埋组织样本来研究NNGR在原发结直肠癌、食道癌和胃癌及其匹配的正常组织中的mRNA和蛋白水平的表达。还将调查NNGR的结构变化，并评估NNGR状态与临床病理特征之间的相关性。还建议进行研究，以阐明NNGR作用的分子机制。为此，将利用诱导系统将外源性NNGR表达到癌细胞中，以系统地研究NNGR蛋白的生物学功能。为了研究NNGR的结构和功能关系，将利用定点突变技术对重要基序进行突变，并研究突变对NNGR生物学和生化功能的影响。将利用Cre/loxP系统有条件地删除小鼠胃肠道中的NNGR，并研究NNGR的缺失对胃肠道形态发生和致癌的影响。这一建议的长期目标是阐明(1)NNGR在胃肠道恶性肿瘤发病机制中的作用；(2)NNGR单独或与其他已知标志物联合作为诊断/预后标志物的地位；(3)确切的分子作用机制。

A novel gene named NNGR (Novel Negative Growth Regulator) has been cloned. NNGR codes for a secreted protein that harbors two zinc finger-like motifs and exhibits homology to yeast killer toxins. NNGR resides at chromosome 4q21.1-22, a region frequently deleted in esophageal and a subset of colorectal cancers. NNGR expression is either lost or decreased in 5/6 colon, 1/1 gastric and 6/7 of esophageal tumors when compared with their matching normal tissues. Most importantly, NNGR expression in NNGR non-expressing colorectal cancer cells inhibits their growth by inducing apoptosis. Thus, NNGR acts as noel growth suppressor whose expression is either lost or decreased in the primary gastrointestinal (GI) cancers. The proposed studies are aimed at the molecular and functional characterization of NNGR. Specifically, fresh frozen and paraffin-embedded tissue specimens will be utilized to investigate the expression of NNGR at the mRNA and protein levels in primary colorectal, esophageal and gastric cancers and matching normal tissues. Structural alterations in NNGR will also be investigated and a correlation between the NNGR-status and clinicopathological features will be evaluated. Studies are also proposed to elucidate the molecular mechanisms of NNGR action. Towards this end, an inducible system will be used to express exogenous NNGR into cancer cells to systematically study the biological function of the NNGR protein. To investigate the structure function relationships, site-directed mutagenesis will be used to mutate important motifs and the effect of mutations on the biological and biochemical functions of NNGR will be investigated. Cre/loxP system will be used to conditionally delete NNGR in the GI tract of mice and the effects of the absence of NNGR on the GI morphogenesis and carcinogenesis will be investigated. The long-term objectives of this proposal are to elucidate (1) the role of NNGR in pathogenesis of the GI malignancies, (2) its status as a diagnostic/prognostic marker either alone or in combination with other known markers and (3) the exact molecular mechanisms of action.