Elucidating the Role of C16orf72 in the Cellular Stress Response Network

项目总结 癌症的一个标志是恶性细胞在面对压力时保持生存的能力，如 非整倍体、营养稀缺和异源化合物。这种多重应激-弹性表型使肿瘤 形成、转移和治疗耐药。癌细胞选择的单个分子通路 促进抗压能力已经得到了彻底的表征，但这些专门的途径是如何 在复杂的生理应激状态下的整合仍然知之甚少。最近，我们使用了基因组规模 适合性筛选数据用于评估689种不同癌细胞的内在应激表型并识别基因 它们在高度依赖于多种不同的应激反应途径的癌细胞中是选择性必不可少的。 通过我们的综合分析，我们发现了C16orf72，一个以前没有特征的蛋白质，它被广泛地 压力感应性，并促进对机械多样的压力侮辱的适应能力。我们已经确定 C16orf72与HUWE1在物理上相互作用，HUWE1是一种E3连接酶，已知泛素化参与应激的蛋白质 反应途径。此外，C16orf72或HUWE1的缺失会产生高度相似的转录和 泛素化表型，而不影响其他因子的表达。因此，我们的首要假设是 C16orf72通过与HUWE1物理相互作用促进HUWE1介导的应激反应而介导应激恢复 在不同的应激反应途径中起关键作用的蛋白质的泛素化。在本提案的目标1中，我们将 定义了C16orf72与HUWE1的绑定接口，测试了C16orf72与HUWE1的几种非独占机型 调节HUWE1的酶活性。在目标2中，我们将确定底物和具体的修饰 C16orf72和HUWE1在细胞应激恢复和典型应激反应信号中的作用。 在目标3中，我们将确定C16orf72在体内肿瘤发生中所需的程度以及 乳腺癌治疗抵抗的研究进展。拟议工作的长期目标是确定 C16orf72促进细胞应激反应的分子机制及治疗研究 阻断癌细胞中C16orf72/HUWE1轴的可能性。

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.