IMMORTALIZATION OF SV40 TRANSFORMED HUMAN CELLS

描述：(改编自申请者的摘要)正常人体细胞 在文化中的寿命有限，并经历复制衰老 而肿瘤细胞通常会持续生长，而且是不朽的。因此 细胞衰老在一定程度上也是一种生长/肿瘤抑制因子 致癌的特定方面。他们一直在剥削 以SV40转化的人成纤维细胞(SV/HF)为模型 永生的分子机制。利用匹配的集合 非永生(永生)和不朽的SV/HF，他们消除了 病毒基因的功能是负责任的，因此试图识别 相关的细胞基因，包括一种名为SEN6的基本初级效应因子 (通过分子遗传学方法)和次级效应器(通过差异 在最后一次授予期间)。每个人 方法导致了一种尚未发现的基因的鉴定 以前被广泛研究或被认为与衰老有关 或者永垂不朽。他们建议调查它们对正常人的影响 Cells和SV/HF。 他们已经确定了一个被灭活的轨迹 永生化，映射到6q27，并在该区域确定了一个基因 它具有生长抑制活性并在不朽的SV/HF中发生突变， 与身为SEN6是一致的。此外，他们还分离出了一种 在不朽的SV/HF中表达降低的基因(与 永生前SV/HF和正常细胞)，其序列表明 蛋白质丝氨酸/苏氨酸磷酸酶(PSP)。拟议的实验将 确定每种药物在SV/HF中是否作为生长抑制因子有效，以及 其他肿瘤来源的永生细胞系，并评估涉及的机制。 SEN6和PSP都具有表明它们可能作用于 共同途径的不同阶段涉及到MAP激酶，因此它们将 最初，他们将注意力集中在永生生物中这些通路的变化上 SV/HF和过表达(在可调控启动子的控制下) SEN6和/或蛋白质候选的引入序列 磷酸酶。他们预计，结果将产生新的见解 细胞衰老及其在癌症发生中的作用。

DESCRIPTION: (Adapted from the applicant's abstract) Normal human cells have a limited lifespan in culture and undergo replicative senescence whereas tumor cells commonly grow continuously and are immortal. Hence cellular senescence also functions in part as growth/tumor suppressor in a particular aspect of carcinogenesis. They have been exploiting SV40-transformed human fibroblasts (SV/HF) as a model to understand the molecular mechanism of immortalization. Utilizing matched sets of non-immortal (preimmortal) and immortal SV/HF, they eliminated changes in viral gene functions as being responsible and, therefore, sought to identify relevant cellular genes including an essential primary effector termed SEN6 (by molecular genetic approaches) and secondary effectors (by differential screening of a cDNA library) in the period of the last grant award. Each approach has resulted in the identification of a gene which had not previously been extensively studied or considered associated with senescence or immortalization. They propose to investigate their effects on normal cells and SV/HF. They have determined that a locus, which is inactivated upon immortalization, maps to 6q27 and have identified a gene in this region which has growth suppressor activity and has been mutated in immortal SV/HF, consistent with being SEN6. In addition, they have isolated a cDNA for a gene whose expression is reduced in immortal SV/HF (as compared to preimmortal SV/HF and normal cells) which has sequences indicative of a protein serine/threonine phosphatase (PSP). Proposed experiments will determine whether each is effective as a growth suppressor in SV/HF and other tumor-derived immortal cell lines, and assess the mechanism involved. Both SEN6 and PSP have properties which indicate that they might act at different stages of a common pathway involving MAP kinases so they will initially focus their attention on changes in those pathways in immortal SV/HF and upon overexpression (under the control of a regulatable promoter) of introduced sequences for the candidate for SEN6 and/or the protein phosphatase. They anticipate that the results will generate new insights into cellular senescence and its role in carcinogenesis.