Global changes in the 3'UTRome toggle responsiveness to growth factors

3UTRome 的整体变化切换对生长因子的反应

• 批准号: 9245281
• 负责人: Marco Mangone
• 金额: $ 19.87万
• 依托单位: TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-10 至 2019-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9245281
• 关键词: 3' Untranslated Regions; Address; Adopted; CRISPR/Cas technology; Caenorhabditis elegans; Catalogs; Cells; Congenital Abnormality; Data Set; Development; Disease; Distal; EGF gene; Epidermal Growth Factor Receptor; Etiology; Event; Extracellular Signal Regulated Kinases; Family; Future; Gene Expression; Generations; Genes; Genetic Translation; Genome engineering; Global Change; Growth Factor; Hallmark Cell; Immunoprecipitation; Lead; Length; MEKs; Malignant Neoplasms; Mammalian Cell; Mediating; Messenger RNA; MicroRNAs; Microarray Analysis; Modality; PIK3CG gene; Pattern; Polyadenylation; Predisposition; Process; Protein Family; RNA-Binding Proteins; Ras/Raf; Regulation; Reporter; Repression; Research; Role; Signal Transduction; Source; Specialized Epithelial Cell; Surveys; System; Testing; Time; Tissues; Untranslated Regions; Variant; Vulva; Work; epigenome; genome editing; mRNA Stability; notch protein; precursor cell; prevent; programs; protein E; response; transcriptome; tumorigenesis

SUMMARY RNA binding proteins like the Pumilio family (PUFs) exert repression on 3'UTRs, and hence mRNA stability and translation. Alternative polyadenylation (APA) causes 3'UTR length to vary. Coordinated large-scale shortening of 3'UTRs through APA can lead to evasion from 3'UTR- mediated repressive signals, and may be a key regulatory regime during development. Yet most identified cases of 3'UTR-mediated PUF repression - and APA to evade repression - were found ad hoc, between interacting gene pairs. Thus a gap exists in our understanding of how these interactions are orchestrated at the global level of the 3'UTRome and APA. EGF patterns the C. elegans vulva. Of six equipotent vulval precursor cells (VPCs), the three closest to the EGF source are induced to form the vulva, while the distal three remain uninduced. Remarkably, this event occurs with 99.8% fidelity. Three redundant PUFs are expressed specifically in the uninduced cells, suggesting that distal VPCs enact a PUF- and 3'UTR-dependent program to become non-responsive to signal. In the germline, the same PUFs repress ERK/MAP kinase; this same mechanism may be adopted by non-responsive VPCs. Germline immunoprecipitation (IP) of one PUF identified many potential target 3'UTRs. From this dataset, we identified multiple target mRNAs from genes in each vulval signaling cascade (EGFR→Ras→Raf→MEK→ERK, Notch→CSL, PI3K→PDK→Akt, and EGFR→Ras→ RalGEF→Ral). We hypothesize that the redundant PUFs collectively repress mRNAs of all four identified signaling cascades to demarcate signal-non-responsive from signal-responsive cells. Our central hypothesis is that switching from proximal (short 3'UTR) to distal (long 3'UTR) polyadenylation sequence (PAS) usage governs switching from signal-responsiveness to non- responsiveness. We will systematically test this hypothesis by joining bottom-up and top-down specific aims. We propose to: 1) test 3'UTRs from the PUF IP list of vulval signaling genes for ability to mediate PUF-dependent reporter repression in non-responsive cells, 2) survey the VPC 3'UTRome and global proximal-to-distal PAS switching, and 3) validate select candidates by altering endogenous 3'UTRs and polyadenylation signals via CRISPR/Cas9 genome editing, and deleting associated PUFs. We will ascertain the contribution to developmental fidelity by APA, and the changes in repressive access points in the 3'UTRs caused by APA.

总结