hnRNP H1 and intronic G-runs in the splicing control of the human rpL3 gene

Alternative splicing (AS) is one of the main regulatory mechanisms of gene expression. It has also been demonstrated that some evolutionarily conserved AS events give rise to aberrant transcripts, that are targeted for decay by nonsense-mediated mRNA (NMD). NMD is an mRNA surveillance pathway that recognizes and selectively degrades mRNAs containing premature termination codons (PTC), thus preventing the synthesis of truncated proteins that could be deleterious to the cell. NMD is also involved, in association with AS, in the post-transcriptional regulation of eukaryotic genes. We previously demonstrated that ASinduced retention of part of intron 3 of rpL3 pre-mRNA produces an mRNA isoform containing a PTC that is substrate of NMD (1). We also demonstrated that overexpression of rpL3 upregulates the AS of its pre-mRNA. Next, we investigated the molecular mechanism underlying rpL3 autoregulation. Specifically we investigated the role played by hnRNP H1 in the regulation of splicing of rpL3 pre-mRNA by manipulating its expression level (2). We also identified and characterized the cisacting regulatory elements involved in hnRNP H1-mediated regulation of rpL3 splicing. RNA electromobility shift assay demonstrated that hnRNP H1 specifically recognizes and binds directly to the intron 3 region that contains seven copies of G-rich elements. Site-directed mutagenesis analysis and in vivo studies showed that the G3 and G6 elements are required for hnRNP H1-mediated regulation of rpL3 pre-mRNA splicing. We propose a working model in which rpL3 recruits hnRNP H1 and, through cooperation with other splicing factors, promotes selection of the alternative splice site. References: 1. Cuccurese M, Russo G, Russo A, Pietropaolo C. Nucleic Acids Res 2005; 33: 5965–5977. 2. Russo A, Siciliano G, Catillo M, Giangrande C, Amoresano A, Pucci P, Pietropaolo C, Russo G. Biochimica et Biophysica Acta 2010;1799(5–6): 419-428