Selective mRNA translation in developmental disorders

摘要 RASopathy引起大多数先天性疾病，影响近1/1000的个体。特别是， RAS-MAPK途径基因中的突变导致不同的病理包括颅面畸形， 精神损伤、肌肉骨骼缺陷和癌症易感性。虽然介绍可能有所不同 在不同的突变之间，几乎所有的RASopathies都有共同的皮肤生长异常。在遗传 在水平上，已知RAS途径成员（包括Hras和Kras）的生殖系突变会导致这些缺陷 其最好的例子是Costello、Noonan和面部皮肤综合征。多年来， RAS在发育过程中的全面研究仅限于DNA的全基因组研究， 核糖核酸这些调查虽然重要，但基本上没有触及预防机制。这是 值得注意的是，新出现的证据表明，发育障碍，如钻石-布莱克凡贫血， 以及Schwachman-Diamond和Treacher柯林斯综合征（综述于Tahmasebi等人，（2018） 与翻译装置的损伤有因果关系。因此，我们目前对机械的认识 RASopathies的基础不完整，这是治疗创新的障碍。我们的长期目标是 揭示Ras介导的组织生长机制，这将最终产生创新疗法， 恢复正常的组织稳态，而不损害发育期间的管家功能。使用 皮肤作为组织发育的确定模型，我们已经发现过度活跃的Hras同时 通过重新连接翻译启动机制来驱动专门的增殖和分化程序 eIF2B5利用我们实验室开创的最先进的体内遗传筛选，我们 确定了eIF 2B 5控制的影响自我更新和细胞命运选择的基因调节子。值得注意的是， 这些mRNA网络通过其功能清楚地划分，其中泛素化作为关键调节因子出现 细胞的分化。因此，我们假设Ras的激活促进了一个子集的翻译。 在发育过程中支持非生理性组织生长的mRNA， 增殖通过它们通过分化成有丝分裂后后代而损失来平衡。在本提案中，我们将 使用体内模型、活体显微镜和新开发的细胞和分子测定的融合 描述RAS和eIF 2B 5之间的相互作用如何影响组织动力学。我们将实现 1）揭示eIF 2B 5依赖的泛素连接酶如何指导祖细胞更新和命运选择; 阐明激活的Hras和eIF 2B 5如何指导mRNA特异性翻译以调节祖细胞 退款总的来说，我们的目标的成功完成将提供一个新的理解蜂窝和 支持Ras驱动的非生理性生长的分子原理。最终，这些 新的见解将为新疗法的发展提供信息，这些疗法可以差异化地抑制病理性 Ras介导组织不平衡的影响，同时维持生命所必需的内稳态。

Abstract RASopathies cause the majority of congenital disorders affecting nearly 1 in 1000 individuals. In particular, mutations in RAS-MAPK pathway genes lead to distinct pathologies including craniofacial dysmorphology, mental impairment, musculoskeletal defects, and a predisposition to cancer. Although presentations may vary between different mutations, nearly all RASopathies share common skin growth abnormalities. At a genetic level, germline mutations to RAS pathway members including Hras and Kras are known to cause these defects which are best exemplified by Costello, Noonan, and Cardiofaciocutaneous syndromes. For years, comprehensive interrogation of RAS in development has been limited to genome-wide studies of DNA and RNA. While important, these investigations have left translation-based mechanisms largely untouched. This is remarkable in light of emerging evidence that developmental disorders, such as Diamond-Blackfan Anemia and Schwachman-Diamond and Treacher Collins syndromes (reviewed in Tahmasebi et al., 2018), are causally linked to impairments in the translation apparatus. Thus, our current knowledge of the mechanistic basis of RASopathies is incomplete, which is a barrier to therapeutic innovation. Our long-term goal is to uncover the mechanism of Ras-mediated tissue growth, which will ultimately yield innovative therapies to restore normal tissue homeostasis without compromising housekeeping functions during development. Using skin as a defined model of tissue development we have discovered that hyperactive Hras simultaneously drives specialized proliferation and differentiation programs by rewiring the translation initiation machinery through eIF2B5. Utilizing state-of-the-art in vivo genetic screens pioneered in our laboratory, we have determined the regulon of genes that eIF2B5 governs to impact self-renewal and cell fate choice. Remarkably, these mRNA networks are clearly demarcated by their function with ubiquitination emerging as a key regulator of cellular differentiation. As such, we hypothesize that activation of Ras promotes translation of a subset of mRNAs that support non-physiological tissue growth during development, where increased stem cell proliferation is balanced by their loss through differentiation into post-mitotic progeny. In this proposal we will use a confluence of in vivo models, intra-vital microscopy, and newly developed cellular and molecular assays to delineate how the interplay between RAS and eIF2B5 influences tissue dynamics. We will accomplish the following Aims: 1) Uncover how eIF2B5-dependent ubiquitin ligases directs progenitor renewal and fate choice; and 2) Elucidate how activated Hras and eIF2B5 direct mRNA specific translation to regulate progenitor renewal. Collectively, the successful completion of our Aims will provide a new understanding of cellular and molecular principles that support Ras-driven non-physiological growth during development. Ultimately, these new insights will inform the development of novel therapeutics, which can differentially inhibit the pathologic impact of Ras mediated tissue imbalance while maintain homeostasis which is essential for life.