The 3’ ends of most eukaryotic mRNAs are cleaved and polyadenylated at the last step of transcription. Recent studies revealed that more than 70% mammalian genes have multiple polyadenylation sites (pAs) leading to the generation of multiple mRNA isoforms with different coding region or 3’ untranslated region (3’ UTR) from the same gene locus and contributes to the complexity of transcriptome and proteome by regulating their stability, localization, translation, and function. Boosted by the large-scale analysis technologies, extensive and dynamic regulation of 3’ UTR by alternative polyadenylation (APA) has been observed in different tissues; different cellular conditions (proliferation, differentiation, and development); and response to stimuli. Although the exact underlying mechanisms of APA remains under investigation, it should be in general regulated via the interaction between cisregulatory elements residing at the DNA/RNA and trans-factors including polyadenylation cleavage core protein complex as well as other accessory RNA binding proteins (RBP). Change of APA pattern during evolution remains underexplored. Such changes could arise from the divergence in cis
-regulatory elements and/or transacting RBPs. The divergences of the two factors with different extent of pleiotropic consequences undergo distinct evolutionary trajectories. Therefore, to better understand evolution in APA, it is important to distinguish the relative contribution of cis
– and trans
-effects. In this project, to comprehensively investigate the contribution of cis
-elements and trans-factors in the process of APA in a mammalian system, we identified and quantified pAs usage difference between two parental strains (C57BL/6J and SPRET/EiJ) and between the two alleles in the F1 hybrids with 3’ read capturing and sequencing (3’ READS) and 3’ mRNA-Seq methods, respectively. In total, we identified 3747 parental divergent pAs across five types of APA, between the two parental mouse strains. By comparing the parental divergent pAs with those in F1 hybrids, we observed a predominant contribution of cis
-regulatory effect on pAs usage, which is mediated by genetic variants between two species around the pAs. Further sequence feature analysis demonstrated that the unstable secondary structure and a novel hexamer UUUUUU in the upstream region of pAs could enhance and inhibit the pAs usage, respectively.
Features QuantSeq 3′ mRNA-Seq Library Prep Kit