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"Regulation and evolution of alternative polyA sites"

by
Zhe Ji
Cell Biology and Molecular Medicine Program
B.S. 2006, Nanjing University



Thesis Advisor: Bin Tian, Ph.D.
Associate Professor
Department of Biochemistry and Molecular Biology

Friday, April 13, 2012
12:00 P.M., MSB-E609


Abstract

The 3’ end cleavage and polyadenylation is an essential step for mRNA maturation. More than half of the protein-coding genes in mammalian genomes have been found to contain multiple polyadenylation sites (polyA sites). Alternative polyadenylation (APA) results in mRNAs with different 3’ untranslated regions (3’UTRs) and/or protein coding sequences (CDS). Since 3’UTRs contain various cis-elements involved in mRNA stability, transport and translation, APA plays important roles in modulation of gene expression. I have applied genomic and bioinformatic techniques to study the regulation of APA in various biological conditions and evolution of polyA sites among species.

First, I report that promoter-distal polyA sites are more used during mouse embryonic development and cell differentiation, resulting in lengthening of 3’UTR. Conversely, 3’UTRs shorten during generation of pluripotent stem cells from somatic cells. Generally, the 3’UTR length is inversely correlated with the cell pluripotent state. The 3’UTR length shows the following trend: germ cells < ES cells < partially differentiated cells < terminally differentiated cells.

Second, using data generated by 3’READS, a high throughput method to map polyA sites and quantify their usage levels, I found that APA affects about 70% of protein coding genes and about 50% of genes expressing long non-coding RNAs (lncRNAs) in the mouse genome. Interestingly, I identified over one thousand polyA sites located in CDS. Using CLIP-seq data for CstF64, a key 3’ end processing factor, I validated these CDS-associated polyA sites. In addition, I found that genomic regions harboring CDS and intronic polyA sites have unique structural features and have fast evolution rates, contributing to species-specific isoform production. Consistently, inhibition of splicing efficiency results in higher usage of CDS polyA sites.

Third, using 3’READS data, I further studied dynamic APA regulation during myogenesis and adipogenesis. I found general lengthening of 3’UTR and more skipping of intronic polyA sites in cell differentiation. Strong polyA sites are more used when cells differentiate. Knockdown of CstF64 leads to a similar global change of APA, but does not fully recapitulate the regulation in cell differentiation, indicating that multiple mechanisms are in play for APA regulation.

Finally, I studied the intestinal gene expression and polyadenylation in C. elegans.


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