Poster abstracts

Poster number 3 submitted by Neha Aggarwal

Nuclear pre-mRNA Splicing and miRNA Processing

Neha Aggarwal (BGES, Cleveland State University), Kavleen Sikand (BGES, Cleveland State University), Girish C. Shukla (BGES, Cleveland State University)

Abstract:
MicroRNAs (miRNAs) are non-coding, endogenous; ~22nt long RNA sequences formed from primary transcripts in two steps by nuclear (Drosha) and cytoplasmic (Dicer) RNases. Many human miRNAs are expressed from introns of protein-coding transcripts, hence we wanted to explore if pre-mRNA splicing and miRNA processing are coupled. We constructed a minigene system with 4 exons and 3 introns from á-MHC gene that harbors miR-208 in intron 28. We induced mutations in the 5’ splice site and the branch site to study the effect of these mutations on pre-mRNA splicing. Additionally, we also incorporated mutations in the upper and lower helical regions of the stem-loop structure of pre-miRNA to study how these mutations affect miRNA processing. In in vivo system, mutations in the pre-miRNA structure had no affect on splicing. However, 5’ splice site mutation activated cryptic splicing. We are using in vitro assay systems to study intermediate splice products generated in both splicing and miRNA processing pathways using the above mentioned mutants.

Keywords: pre-mRNA splicing, Micro RNA processing