title: G-quadruplex formation enhances splicing efficiency of PAX9 intron 1.
abstract: G-quadruplexes are secondary structures present in DNA and RNA molecules, which are formed by stacking of G-quartets (i.e., interaction of four guanines (G-tracts) bounded by Hoogsteen hydrogen bonding). Human PAX9 intron 1 has a putative G-quadruplex-forming region located near exon 1, which is present in all known sequenced placental mammals. Using circular dichroism (CD) analysis and CD melting, we showed that these sequences are able to form highly stable quadruplex structures. Due to the proximity of the quadruplex structure to exon-intron boundary, we used a validated double-reporter splicing assay and qPCR to analyze its role on splicing efficiency. The human quadruplex was shown to have a key role on splicing efficiency of PAX9 intron 1, as a mutation that abolished quadruplex formation decreased dramatically the splicing efficiency of human PAX9 intron 1. The less stable, rat quadruplex had a less efficient splicing when compared to human sequences. Additionally, the treatment with 360A, a strong ligand that stabilizes quadruplex structures, further increased splicing efficiency of human PAX9 intron 1. Altogether, these results provide evidences that G-quadruplex structures are involved in splicing efficiency of PAX9 intron 1.
authors.affiliation: Department of Morphology, Piracicaba Dental School, University of Campinas-UNICAMP, Piracicaba, SP, 13414-903, Brazil, maririb@gmail.com.
authors.forename: Mariana Martins
authors.initials: MM
authors.lastname: Ribeiro
authors.forename: Gleidson Silva
authors.initials: GS
authors.lastname: Teixeira
authors.forename: Luciane
authors.initials: L
authors.lastname: Martins
authors.forename: Marcelo Rocha
authors.initials: MR
authors.lastname: Marques
authors.forename: Ana Paula
authors.initials: AP
authors.lastname: de Souza
authors.forename: Sergio Roberto Peres
authors.initials: SR
authors.lastname: Line
mesh_terms: D000818:Animals; D001483:Base Sequence; D002942:Circular Dichroism; D031541:Fluorescence Resonance Energy Transfer; D054856:G-Quadruplexes; D006801:Humans; D007438:Introns / Q000235:genetics*; D008969:Molecular Sequence Data; D051764:PAX9 Transcription Factor / Q000737:chemistry* / Q000235:genetics*; D012326:RNA Splicing / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012689:Sequence Homology, Nucleic Acid
chemical_list: D051764:PAX9 Transcription Factor; C497323:PAX9 protein, human; D012333:RNA, Messenger