Functional characterization of alternatively spliced variants of human FOXP2 speech gene.

Abstract

FOXP2 gene encodes a forkhead transcription factor recently linked to autism spectrum disorders (ASDs). Three major FOXP2 spliced variants have been described: a full-length (FOXP2-FL), a shorter transcript starting at exon 4 (FOXP2-III) and an isoform truncated at exon 10 (FOXP2-10+). FOXP2-10+ mRNA has a short 3'-UTR and encodes a protein lacking the DNA binding domain, whose transient expression was detected during mouse brain development. This study aims to investigate the functional role of FOXP2 variants in cellular models and characterize the RNA-binding proteins involved in transcript maturation. In Hek293 cells, of the three co-expressed transcripts, only FOXP2-FL and FOXP2-III were translated into peptides, questioning the functional role of FOXP2-10+ transcript. In silico analysis of FOXP2 gene revealed clusters of putative binding sites for Polypyrimidine-Tract Binding protein 1 (PTBP1), a regulator of exon-skipping events in tissue differentiation, including neuronal development. PTBP1 consensus sites were located in intron sequences flanking two exons of FOXP2 and, interestingly, within the 3'-UTR of FOXP2-FL. Based on these findings, we overexpressed PTBP1 in Hek293 cells and analyzed FOXP2 transcript profile. By qPCR, no significant change was observed in ratios between FOXP2 variants, but we detected a novel transcript excluding exon 11 (FOXP2-\u39411), lacking a nuclear localization signal sequence. Of note, intron sequences flanking exon 11 contain putative PTBP1 binding sites. Currently, we are investigating FOXP2 transcript expression upon silencing of endogenous PTBP1 in Hek293 cells and during mouse brain development. We propose PTBP1 as a regulator of FOXP2 transcripts maturation or stability