Thousands of human genes produce multiple distinct mRNA isoforms through alternative cleavage and polyadenylation (APA). APA is often conserved in other mammals. It can dramatically change protein function, e.g., switching between membrane-bound and secreted forms. It can sometimes produce mRNA isoforms that differ in their stability or translation, but we have found that this is uncommon. Instead, we hypothesize that APA is often regulated for the purpose of producing mRNA isoforms that differ in their subcellular localization, such as localization to axons and dendrites of neurons. Our approach to the long-term goal of understanding the regulation and function of APA in mammals is organized around the following specific aims: SA1. Determine the dynamics and function of mRNA 3' ends in neuronal differentiation. We propose to map the dynamics of alternative 3' UTR expression during neuronal differentiation in vitro and to assess the localization properties of alternative 3' UTRs. SA2. Identify factors that control the neuronal program of alternative 3' UTR isoforms. We will integrate data from SA1 and from an ENCODE project related to RNA binding proteins to identify candidate APA regulatory factors, and will then test them by analysis of mRNA isoforms following RNAi, and by use of a metabolic labeling approach to distinguish regulation of cleavage and polyadenylation (CPA) from regulation of mRNA stability. Fundamentally, this proposal seeks to understand the regulation and function of the 3' ends of genes, and to establish their roles in the nervous system, with potential implications for neurological disease and cancer.

Public Health Relevance

This project will describe changes to the patterns of transcript expression that occur as precursor cells become neurons, and provide insights into the functions of these transcripts and identify proteins that regulate these patterns. The patterns of transcript expression that occur in neurons are often reversed in cancer, and this study may provide clues as to how to prevent cancer cells from acquiring their characteristic transcript expression patterns.

Agency
National Institute of Health (NIH)
Institute
National Human Genome Research Institute (NHGRI)
Type
Research Project (R01)
Project #
5R01HG002439-15
Application #
9272911
Study Section
Genomics, Computational Biology and Technology Study Section (GCAT)
Program Officer
Gilchrist, Daniel A
Project Start
2002-02-06
Project End
2019-05-31
Budget Start
2017-06-01
Budget End
2018-05-31
Support Year
15
Fiscal Year
2017
Total Cost
$489,006
Indirect Cost
$172,780
Name
Massachusetts Institute of Technology
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
001425594
City
Cambridge
State
MA
Country
United States
Zip Code
02142
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Taliaferro, J Matthew; Lambert, Nicole J; Sudmant, Peter H et al. (2016) RNA Sequence Context Effects Measured In Vitro Predict In Vivo Protein Binding and Regulation. Mol Cell 64:294-306
Merkin, Jason J; Chen, Ping; Alexis, Maria S et al. (2015) Origins and impacts of new mammalian exons. Cell Rep 10:1992-2005
Katz, Yarden; Wang, Eric T; Silterra, Jacob et al. (2015) Quantitative visualization of alternative exon expression from RNA-seq data. Bioinformatics 31:2400-2
Lambert, Nicole; Robertson, Alex; Jangi, Mohini et al. (2014) RNA Bind-n-Seq: quantitative assessment of the sequence and structural binding specificity of RNA binding proteins. Mol Cell 54:887-900
Shalgi, Reut; Hurt, Jessica A; Krykbaeva, Irina et al. (2013) Widespread regulation of translation by elongation pausing in heat shock. Mol Cell 49:439-52
Spies, Noah; Burge, Christopher B; Bartel, David P (2013) 3' UTR-isoform choice has limited influence on the stability and translational efficiency of most mRNAs in mouse fibroblasts. Genome Res 23:2078-90
Han, Hong; Irimia, Manuel; Ross, P Joel et al. (2013) MBNL proteins repress ES-cell-specific alternative splicing and reprogramming. Nature 498:241-5
Hurt, Jessica A; Robertson, Alex D; Burge, Christopher B (2013) Global analyses of UPF1 binding and function reveal expanded scope of nonsense-mediated mRNA decay. Genome Res 23:1636-50
Klattenhoff, Carla A; Scheuermann, Johanna C; Surface, Lauren E et al. (2013) Braveheart, a long noncoding RNA required for cardiovascular lineage commitment. Cell 152:570-83

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