HnRNPA1 interacts with G-quadruplex in the TRA2B promoter and stimulates its transcription in human colon cancer cells

The human TRA2B gene consists of 10 exons and 9 introns and produces 5 splice isoforms (TRA2 beta 1 to TRA2 beta 5). TRA2B exon 2 encodes multiple premature termination codons. TRA2 beta 1 lacks exon 2 and is translated into a functional transformer 2 beta (Tra2 beta) protein, whereas TRA2 beta 4 contains 10 exons and works as a functional RNA. Overexpressed Tra2 beta and ectopic expression of TRA2 beta 4 may be oncogenic. We found that heterogeneous nuclear ribonucleoprotein (hnRNP) A1 and hnRNPU interacted with TRA2 beta 4 exon 2. Minigene assays revealed that hnRNPA1 facilitated inclusion of exon 2, whereas hnRNPU promoted its skipping. However, knockdown of hnRNPA1 or hnRNPU reduced both TRA2 beta 1 and TRA2 beta 4 levels, and overexpression of these hnRNPs increased levels of both isoforms, suggesting that hnRNPA1 and hnRNPU mainly regulate the transcription of TRA2B. In fact, hnRNPA1 and hnRNPU positively regulated the promoter activity of TRA2B. Circular dichroism analyses, electrophoretic mobility shift assays and chromatin immunoprecipitation assays demonstrated the presence of G-quadruplex (G4) formation in the promoter of TRA2B. Formation of G4 suppressed TRA2B transcription, whereas hnRNPA1, but not hnRNPU, interacted with the G4 to facilitate transcription. Our results suggest that hnRNPA1 may modulate TRA2B transcription through its regulation of G4 formation in its promoter in colon cancer cells.