hnRNP A1 controls MIR17HG alternative splicing and maturation of miR-19a

Abstract

Alternative splicing is a key process in the regulation of gene expression in eukaryotes. During transcription, the spliceosome machinery assembles onto precursor messenger RNA (pre-mRNA) conserved regions, promoting intron removal and exon ligation. This process is guided and controlled by the association of RNA-binding proteins (RBPs) in the pre-mRNAs. These proteins mediate splice-site choice controlling the splicing profile observed in different cells. The heterogeneous ribonucleoproteins (hnRNPs) A1 and K have previously been associated with intronic regions containing microRNAs (miRNAs). We previously showed hnRNP A1 overexpression increases invasion and cell growth in thyroid carcinoma cells through association with miR-17a and miR-18a. MIR17HG is the host gene for the miR-17-92 miRNA cluster, which is encoded within one of its introns. In this work, we investigated the participation of hnRNP A1 and hnRNP K in MIR17HG alternative splicing. Overexpression of hnRNP A1 and hnRNP K was performed using pFLAG system in HEK-293FT cells. Reverse transcription quantitative PCR (RT-qPCR) analysis indicated higher levels of MIR17HG-i3 isoforms, which correlated with increase on pre-miR-19a generation upon hnRNP A1 overexpression. Surprisingly, the same pattern was not observed in cells overexpressing hnRNP K. Higher concentration of functional miR-19a in hnRNP A1 overexpressing cells was confirmed by luciferase reporter assays. These results indicate that hnRNP A1 modulates MIR17HG splicing, with downstream effects on miR-19a biogenesis and maturation.