Characterization of a Novel Growth Regulator

NNGR（Novel Negative Growth Regulator）是一种新的负性生长调节因子。NNGR编码一种分泌蛋白，具有两个锌指样基序，与酵母杀手毒素具有同源性。NNGR位于染色体4q21.1-22，这是食管癌和结肠直肠癌的一个子集中经常缺失的区域。与相应的正常组织相比，5/6的结肠癌、1/1的胃癌和6/7的食管癌中NNGR表达缺失或降低。最重要的是，NNGR非表达结直肠癌细胞中的NNGR表达通过诱导细胞凋亡来抑制其生长。因此，NNGR作为诺埃尔生长抑制因子，其在原发性胃肠道（GI）癌中表达缺失或降低。拟议的研究旨在NNGR的分子和功能表征。具体而言，将使用新鲜冷冻和石蜡包埋的组织标本来研究原发性结直肠癌、食管癌和胃癌以及匹配的正常组织中NNGR在mRNA和蛋白水平上的表达。还将研究NNGR的结构改变，并评价NNGR状态与临床病理学特征之间的相关性。研究还提出了阐明NNGR行动的分子机制。为此，将使用诱导系统将外源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的结构和功能之间的关系，将采用定点突变来突变重要的基序，并研究突变对NNGR的生物学和生化功能的影响。将使用Cre/loxP系统来条件性地删除小鼠胃肠道中的NNGR，并且将研究NNGR的缺失对胃肠道形态发生和癌变的影响。本提案的长期目标是阐明（1）NNGR在GI恶性肿瘤发病机制中的作用，（2）其作为单独或与其他已知标志物联合的诊断/预后标志物的地位，以及（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.