Mechanisms of Induction and Suppression of Viral and Tum

我们一直在研究通过丝裂原活化蛋白激酶（MAPK）信号通路起作用的癌基因的转化机制。Drm（Down-regulated by mos）/Gremlin是一种骨形态发生蛋白（BMP）拮抗剂，最初由我们实验室鉴定，在体内以组织特异性方式表达。除原代成纤维细胞和少数细胞系外，培养中的大多数转化细胞不能表达DRM/Gremlin，我们观察到其表达可以通过MAPK途径被癌基因转化抑制。我们假设这种表达缺失可能对特定肿瘤的发生或进展很重要。我们最近的工作集中在分析Drm在转化细胞中的功能和作用机制。 当将表达DRM/Gremlin的逆转录病毒和质粒克隆引入表达很少或不表达Drm/Gremlin的转化细胞中时，一些细胞显示出生长和转化相关表型特征的改变。为了更仔细地检查Drm/Gremlin表达对肿瘤细胞系的影响，我们构建了蜕皮激素诱导的Drm/Gremlin构建体，并在用诱导剂Ponasterone A处理后产生过表达Drm/Gremlin的细胞系。同时，我们产生了组成型过表达蛋白质的细胞系。当Drm/Gremlin过表达时，p53阴性的原始神经外胚层来源的Daoy和骨肉瘤来源的Saos 2细胞系在培养中表现出较慢的生长，并且体内肿瘤形成能力降低。相反，几种p53表达细胞系显示出增加的生长和肿瘤形成能力，或与对照细胞相比没有变化。在Daoy和Saos 2中，Drm/Gremlin的过表达与p21 Cip 1/Waf 1的过表达相关，p21 Cip 1/Waf 1是参与G1/S细胞周期转换的细胞周期蛋白依赖性激酶的已知抑制剂。其他生长相关蛋白，包括ERK 1/ERK 2的水平没有受到影响。初步证据表明，p21启动子活性降低后，Drm/Gremlin诱导这些细胞。相比之下，HT 1080（纤维肉瘤）细胞表现出p21 Cip 1/Waf 1水平降低，增殖和致瘤率增加。这些数据表明，BMP拮抗剂Drm/Gremlin可以以新的方式影响肿瘤细胞的生长和功能。

We have been studying the transformation mechanisms of oncogenes that act through the Mitogen Activated Protein Kinase (MAPK) signaling pathway. Drm (Down-regulated by mos)/Gremlin is a Bone Morphogenetic Protein (BMP) antagonist, initially identified by our lab, which is expressed in a tissue-specific fashion in vivo. With the exception of primary fibroblasts and a few cell lines, most transformed cells in culture fail to express DRM/Gremlin, and we observed that its expression could be suppressed by oncogene transformation, via the MAPK pathway. We hypothesized this loss of expression may be important for the initiation or progression of specific tumors. Our recent work has focused on analyzing Drm's function and mechanism of action in transformed cells. When retroviral and plasmid clones expressing DRM/Gremlin were introduced into transformed cells that expressed little or no Drm/Gremlin, some showed alterations in growth and transformation-associated phenotypic characteristics. To more carefully examine the effect of Drm/Gremlin expression on tumor cell lines, we constructed an ecdysone-inducible Drm/Gremlin construct and generated cell lines that overexpressed Drm/Gremlin following treatment with the inducer Ponasterone A. In parallel we generated cell lines which constitutively overexpress the protein. The p53-negative, primitive neuroectodermal-derived Daoy and the osteosarcoma-derived Saos2 cell lines exhibited slower growth in culture and reduced tumor-forming ability in vivo when Drm/Gremlin was overexpressed. In contrast several p53-expressing cell lines show either increased growth and tumor forming ability, or no change from control cells. In Daoy and Saos2, overexpression of Drm/Gremlin correlated with overexpression of p21Cip1/Waf1, a known inhibitor of the cyclin-dependent kinases involved in progression through the G1/S cell cycle transition. Levels of other growth associated proteins, including ERK1/ERK2, were not affected. Preliminary evidence suggests that p21 promoter activity is decreased following Drm/Gremlin induction in these cells. In contrast, HT1080 (fibrosarcoma) cells exhibit a reduction in p21Cip1/Waf1 levels and increased rates of proliferation and tumoriginicity. These data suggest that the BMP antagonist Drm/Gremlin can act in novel ways to affect tumor cell growth and function.