PTBP1 promotes the alternative splicing of autism-associated FOXP2 gene

Forkhead Box P2 (FOXP2) gene, implicated in speech and language disorders, encodes a transcriptional regulator of neuronal development recently associated with autism spectrum disorders (ASDs), although controversially. Indeed, no mutations have been identified in FOXP2 coding regions of ASDs patients, thus pointing to a possible altered post-transcriptional control of FOXP2 mRNAs. Our study focuses on the regulation of FOXP2 alternatively spliced mRNAs and the ribonucleoproteins involved. FOXP2 full-length (FOXP2-FL) coding transcript is formed by 17 exons and two rarely in frame included short exons, 3b and 4a. In silico analyses of FOXP2 sequences using SFmap software identified, in introns flanking exons 3b and 11, putative binding sites for PTBP1 (Polypyrimidine Tract Binding Protein 1), a key player in neuronal differentiation. Under these premises, we overexpressed PTBP1 in HEK293 cells and found that PTBP1 significantly down-regulated FOXP2 expression at both mRNA and protein levels. Interestingly, the reduction of FOXP2-FL mRNA appeared concomitant with the detection of increased levels of a novel FOXP2 transcript lacking exon 11 (FOXP2-11). Exon 11-exclusion introduces a premature stop codon (PTC) within exon 12, which will generate a truncated FOXP2 protein lacking the DNA binding domain, if translated. We could not find FOXP2-11-derived peptides, but FOXP2-11 transcripts were detected in several other cell lines expressing FOXP2 and PTBP1. A FOXP2 minigene assay further confirmed PTBP1 ability to promote the skipping of exon 11. With similar approaches, we found that PTBP1 does not regulate FOXP2 exon 3b, suggesting specificity towards exon 11. Finally, by using selective inhibitors, we showed that FOXP2-11 transcript is a target of Nonsense-Mediated Decay (NMD), a conserved degradation pathway for aberrant or PTC-bearing mRNAs. All together, our results show that PTBP1 may control FOXP2 protein expression by increasing the levels of a newly described non-coding FOXP2-11 transcript at the expenses of FOXP2-FL mRNA.