Gadd 45 genes sensors of stress and tumor modulators

描述(申请人提供)：GADD45基因(a，b，g)是压力感受器，调节细胞对遗传毒性/生理压力的反应，并调节肿瘤的形成。GADD45蛋白与其他应激反应蛋白相互作用，包括增殖细胞核抗原、p21、CDc2/Cylin B1、MEKK4和p38激酶。GADD45蛋白的功能在多大程度上重叠，以及应激刺激的性质如何决定GADD45功能以传递细胞存活或细胞死亡的信号，目前尚不清楚。测试的假设是，压力的性质/大小决定了GADD45蛋白将与哪些伙伴相关联，以通知细胞生存或细胞死亡。在低应激条件下，GADD45蛋白可能与p21、cdc2/cyClinB1和增殖细胞核抗原相互作用，激活细胞周期停滞和DNA修复，促进细胞存活；而在高应激条件下，包括细胞老化和癌基因激活，GADD45蛋白可能与应激蛋白(MEKK4、p38、JNK)相互作用，促进细胞凋亡或衰老。推测GADD45蛋白的应激敏感功能在调节肿瘤的形成中发挥作用。缺乏一个或多个GADD45基因的小鼠&缺乏与特定伴侣结合的GADD45突变蛋白来检验这一假设。目的1将评估GADD45及其相互作用伙伴在细胞对不同剂量的遗传毒性应激反应中的作用。GADD45缺陷对GADD45 KO和WT细胞的细胞周期停滞、存活或凋亡的影响，将在体外和体内，暴露于低水平/高水平的基因毒性应激后进行测试。此外，还将通过测试转导的wt/突变体GADD45基因拯救wt表型的能力来探索GADD45/伙伴蛋白相互作用的作用。目的2将评估GADD45及其伙伴在MEF对生理/致癌应激反应中的作用。Gadd45a-/-和Gadd45G-/-MEF可逃脱复制性和癌基因介导的衰老，而Gadd45b-/-MEF则丧失活性。Gadd45a-/-&gadd45G-/-MEF也对ras转化敏感，而gadd45a/gadd45G双KO MEF对ras或myc转化敏感。GADD45和相互作用的伙伴在衰老、存活和对转化的敏感性中所起的作用将被探索。目的3将评估GADD45基因作为肿瘤发展调节因子的功能。产生乳腺癌易感的MMTV-ras和MMTV-myc小鼠wt或gadd45a为空。Gadd45a缺乏促进了MMTV-ras肿瘤的形成，但抑制了MMTV-myc的癌变，表明Gadd45a依赖于癌基因发挥肿瘤的促进/抑制作用。实验的目的是评估GADD45基因的应激反应功能在差异调控ras/myc驱动的乳腺肿瘤发生中所起的作用。获得的数据将对更好地理解应力传感器在肿瘤发生中的作用以及癌症治疗中治疗抵抗如何产生和消除具有重要意义。

DESCRIPTION (provided by applicant): Gadd45 genes (a, b, g) are stress sensors that modulate the response of cells to genotoxic/physiological stress, & modulate tumor formation. Gadd45 proteins interact with other stress response proteins, including PCNA, p21, Cdc2/CyclinB1, MEKK4 & p38 kinase. To what extent the functions of Gadd45 proteins overlap, & how the nature of stress stimuli dictate Gadd45 functions to signal cell survival or cell death is unclear. The hypothesis tested is that the nature/magnitude of stress dictates which partners Gadd45 proteins will associate with to signal cell survival or cell death. In response to low stress Gadd45 proteins may interact with p21, cdc2/cyclinB1 & PCNA to activate cell cycle arrest and DNA repair to promote cell survival, whereas in response to high stress, including cellular aging & activated oncogenes, Gadd45 proteins may interact with stress kinases (MEKK4, p38, JNK) to promote apoptosis or senescence. It is surmised that stress sensing functions of Gadd45 proteins play a role in modulating tumor formation. Mice deficient for one or more gadd45 genes & Gadd45 mutant proteins deficient in binding to particular partners were generated to test the hypothesis. Aim 1 will assess the role of Gadd45 & interacting partners in the response of cells to varying doses of genotoxic stress. The effect of Gadd45 deficiencies on cell cycle arrest, survival or apoptosis in gadd45 KO & WT cells in vitro & in vivo, following exposure to low/high levels of genotoxic stress will be tested. Also, the role of Gadd45/partner protein interactions will be explored by testing the ability of transduced wt/mutant gadd45 genes to rescue wt phenotypes. Aim 2 will assess role of Gadd45 & partners in the response of MEFs to physiological/oncogenic stress. Gadd45a-/- & gadd45g-/- MEFs were found to escape replicative & oncogene mediated senescence, whereas gadd45b-/- MEFs lose viability. Gadd45a-/- & gadd45g-/- MEFs were also found to be susceptible to ras transformation, whereas gadd45a/gadd45g double KO MEFs were susceptible for transformation by ras or myc. The role Gadd45 & interacting partners play in senescence, survival & susceptibility to transformation will be explored. Aim 3 will assess function of gadd45 genes as modulators of tumor development. Breast cancer prone MMTV-ras & MMTV-myc mice wt or null for gadd45a were generated. Gadd45a deficiency accelerated MMTV-ras tumor formation, yet retarded MMTV-myc carcinogenesis, indicating that gadd45a functions as tumor promoter/suppressor depending on the oncogene. Experiments are targeted at assessing the role stress response functions of gadd45 genes play in differentially modulating ras/myc driven breast tumorigenesis. Data obtained will be of great importance to better understand the role of stress sensors in tumorigenesis, & how treatment resistance in cancer therapy can arise & be abrogated.