GENETIC ANALYSIS OF RAS TRANSFORMATION AND JUN FUNCTION

描述：（改编自申请人摘要）： ras原癌基因的激活和转录激活， c-Jun和AP-1转录因子的反式激活。 申请人 在小鼠体内制造了c-jun的无效突变 原代细胞 从这样的胚胎在培养物中生长立即停止。 生长停滞可以 用大T抗原和活化Ras共转化克服; 然而，与由相同试剂转化的野生型细胞不同， 保持非致瘤性。 这些数据表明c-jun在Ras中的关键作用 诱导肿瘤形成。 申请人建议进一步阐明这一点， 使用生物化学和遗传学相结合的方法。 首先，他会 用S63/S73突变体靶向替换c-jun，以评估其作用 JNK介导的磷酸化对c-jun功能的影响。 类似 将采用各种方法来确定 δ区，被认为在JNK的募集和 c-jun的泛素化/不稳定化。 不同的c-jun基因将通过c-jun被替换的研究来评估 由Jun B或Jun D。此外，申请人将评估junD的作用 直接将他击倒 最后，申请人将使用 肿瘤发生建立和候选物筛选的新体内测定法 ras/AP-1介导肿瘤发生的靶基因。

DESCRIPTION: (adapted from applicant's abstract): A critical link between the activation of the ras proto-oncogene and transcriptional activation is the transactivation of c-Jun and AP-1 transcription factors. The applicant has created a null mutation of c-jun in the mouse. Primary cells derived from such embryos growth arrest immediately in culture. Growth arrest can be overcome by co-transformation with large T antigen and activated Ras; however, these cells, unlike wild type cells transformed by the same agents, remain non-tumorigenic. These data indicate a critical role of c-jun in Ras induced tumor formation. The applicant proposes to further elucidate this role using a combined biochemical and genetic approach. First, he will perform targeted replacement of c-jun with a S63/S73 mutant, to assess role of JNK-mediated phosphorylation on c-jun function in vivo. Similar approaches will be used to determine the role of and the requirement for the delta region, which is thought to play a role both in JNK recruitment and in ubiquitination/destabilization of c-jun. The possible redundancy of different c-jun genes will be assessed by studies in which c-jun is replaced by jun B or jun D. In addition, the applicant will assess the role of junD directly by performing a junD knockout. Finally, the applicant will use a novel in vivo assay of tumorigenic establishment and screen for candidate target genes involved in ras/AP-1 mediate tumorigenesis.