Elucidating the Role of C16orf72 in the Cellular Stress Response Network

项目摘要 癌症的一个标志是恶性细胞在面对压力源时保持活力的能力， 非整倍体、营养缺乏和异生化合物。这种多应激-恢复表型使肿瘤 形成、转移和治疗抗性。癌细胞选择的单个分子通路 促进压力恢复能力已经彻底的特点，但这些专门的途径是如何 在复杂的生理应激状态中的整合仍然知之甚少。最近，我们使用基因组规模 适合度筛选数据，以估计689种不同癌细胞的内在应力表型并鉴定基因 其在高度依赖于多种不同应激反应途径的癌细胞中是选择性必需的。 通过我们的综合分析，我们发现了C16 orf 72，一种以前未表征的蛋白质， 压力诱导型，并促进对机械上不同的压力损伤的恢复力。我们已经确定 C16 orf 72与HUWE 1物理相互作用，HUWE 1是一种已知参与应激的泛素化蛋白质的E3连接酶 响应路径。此外，C16 orf 72或HUWE 1的缺失产生高度相似的转录组学和 泛素化表型，而不影响其他因子的表达。因此，我们的总体假设是 C16 orf 72通过与HUWE 1物理相互作用来介导应激恢复，从而促进HUWE 1介导的应激恢复。 泛素化的蛋白质在不同的压力反应途径的关键作用。在本建议的目标1中，我们将 定义了C16 orf 72与HUWE 1的绑定接口，并测试了C16 orf 72与HUWE 1的几个非独占模型， 调节HUWE 1酶活性。在目标2中，我们将确定底物和特定修饰， C16 orf 72和HUWE 1在细胞应激恢复和典型应激反应信号传导中的作用。 在目标3中，我们将确定C16 orf 72在体内肿瘤发生中所需的程度，以及C16 orf 72在体内肿瘤发生中的作用。 乳腺癌治疗耐药性的发展。拟议工作的长期目标是确定 C16 orf 72促进细胞应激恢复的分子机制，并研究治疗 在癌细胞中阻断C16 orf 72/HUWE 1轴的潜力。

PROJECT SUMMARY A hallmark of cancer is the ability of malignant cells to maintain viability in the face of stressors such as aneuploidy, nutrient scarcity, and xenobiotic compounds. This multi-stress-resilience phenotype enables tumor formation, metastasis, and therapy resistance. Individual molecular pathways co-opted by cancer cells to promote stress resilience have been thoroughly characterized, but how these specialized pathways are integrated in complex physiological stress states remains poorly understood. Recently, we used genome-scale fitness screening data to estimate the intrinsic stress phenotype of 689 diverse cancer cell and identify genes which are selectively essential in cancer cells highly reliant on multiple distinct stress response pathways. Through our integrative analysis, we discovered C16orf72, a previously uncharacterized protein which is broadly stress-inducible and promotes resilience to mechanistically diverse stress insults. We have determined that C16orf72 physically interacts with HUWE1, an E3 ligase known to ubiquitinate proteins involved in stress response pathways. Moreover, loss of either C16orf72 or HUWE1 produces highly similar transcriptomic and ubiquitination phenotypes while not affecting expression of the other factor. Thus, our overarching hypothesis is that C16orf72 mediates stress resilience by physically interacting with HUWE1 to promote HUWE1-mediated ubiquitination of proteins with critical roles in diverse stress response pathways. In Aim 1 of this proposal, we will define the binding interface of C16orf72 and HUWE1 and test several non-exclusive models by which C16orf72 regulates HUWE1 enzymatic activity. In Aim 2, we will determine the substrates and specific modifications which underly the role of C16orf72 and HUWE1 in cellular stress resilience and canonical stress response signaling. In Aim 3, we will determine the extent to which C16orf72 is required for in vivo tumorigenesis and the development of therapy resistance in breast cancer. The long-term goals of the proposed work are to determine the molecular mechanism by which C16orf72 promotes cellular stress resilience and investigate the therapeutic potential of blocking the C16orf72/HUWE1 axis in cancer cells.