TELOMERASE ASSOCIATED FACTORS AND THEIR ROLES IN CANCER PROGRESSION

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。癌细胞通常上调核糖核蛋白端粒酶。先前预测端粒酶的抑制可以减缓癌症的生长，但只有在端粒缩短的足够时间过去后才会发生。最近，我们通过一种靶向野生型端粒酶RNA模板区的siRNA特异性抑制端粒酶功能，从而抑制了癌细胞的生长。这种siRNA在癌细胞中的表达导致端粒酶活性急剧降低，细胞生长迅速受到抑制，并诱导细胞凋亡。出乎意料的是，这些有效的细胞效应并不是由端粒结构的破坏引起的，这表明siRNA对端粒酶的抑制可能抑制了端粒酶在癌细胞中的功能，而不是端粒长度的维持。这些细胞中的DNA微阵列分析表明，生长相关基因表达的快速变化不依赖于双链RNA引发的干扰素反应。这些数据表明，除了端粒长度维持外，端粒酶可能对癌细胞增殖至关重要。因此，我们的主要研究重点是通过鉴定新的端粒酶相关因子来进一步了解端粒酶调节细胞增殖的潜在机制。采用串联亲和纯化（TAP）技术对人癌细胞Hela S3的端粒酶复合物进行生化纯化。与端粒酶特异性相关的蛋白质将通过质谱法分离和鉴定。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Cancer cells commonly upregulate the ribonucleoprotein-telomerase. Inhibition of telomerase has previously been predicted to slow cancer growth, but only after a sufficient period elapses for telomere shortening to occur. Recently, we have inhibited cancer cell growth specifically by inhibiting telomerase function through an siRNA targeting the template region of wild-type telomerase RNA. Expression of such siRNA in cancer cells resulted in dramatic reduction of telomerase activity, rapid inhibition of cell growth and induction of apoptosis. Unexpectedly, those potent cellular effects were not apparently induced by the disruption of telomere structures, suggesting that inhibition of telomerase by siRNA may inhibit a function of telomerase in cancer cells other than telomere length maintenance. DNA microarray analysis in these cells indicates rapid changes in growth-related gene expression that is independent of interferon response triggered by double stranded RNA. These data suggested that telomerase may be essential for cancer cell proliferation in addition to telomere length maintenance. Therefore, a main focus of our research is to further understand the underlying mechanisms by which telomerase regulates cell proliferation through identification of novel telomerase associated factors. Biochemistry purification of telomerase complexes from human cancer cell line Hela S3 has been achieved using Tandem affinity purification (TAP) protocol. Proteins associated specifically with telomerase will be isolated and identified by Mass Spectrometry.