Depletion of TDP 43 overrides the need for exonic and intronic splicing enhancers in the human apoA-II gene.
Exon 3 of the human apolipoprotein A-II (apoA-II) gene is efficiently included in the mRNA although its acceptor site is significantly weak because of a peculiar (GU)16 tract instead of a canonical polypyrimidine tract within the intron 2/exon 3 junction. Our previous studies demonstrated that the SR proteins ASF/SF2 and SC35 bind specifically an exonic splicing enhancer (ESE) within exon 3 and promote exon 3 splicing. In the present study, we show that the ESE is necessary only in the proper context. In addition, we have characterized two novel sequences in the flanking introns that modulate apoA-II exon 3 splicing. There is a G-rich element in intron 2 that interacts with hnRNPH1 and inhibits exon 3 splicing. The second is a purine rich region in intron 3 that binds SRp40 and SRp55 and promotes exon 3 inclusion in mRNA. We have also found that the (GU) repeats in the apoA-II context bind the splicing factor TDP-43 and interfere with exon 3 definition. Significantly, blocking of TDP-43 expression by small interfering RNA overrides the need for all the other cis-acting elements making exon 3 inclusion constitutive even in the presence of disrupted exonic and intronic enhancers. Altogether, our results suggest that exonic and intronic enhancers have evolved to balance the negative effects of the two silencers located in intron 2 and hence rescue the constitutive exon 3 inclusion in apoA-II mRNA.
Mercado
Pablo Arrisi
PA
International Centre for Genetic Engineering and Biotechnology, Padriciano 99, I-34012 Trieste, Italy.
Ayala
Youhna M
YM
Romano
Maurizio
M
Buratti
Emanuele
E
Baralle
Francisco E
FE
D016633:Apolipoprotein A-II / Q000235:genetics* / Q000378:metabolism; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D005091:Exons; D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H / Q000378:metabolism; D006801:Humans; D007438:Introns; D009687:Nuclear Proteins / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D034622:RNA Interference; D012326:RNA Splicing; D016601:RNA-Binding Proteins; D038621:Regulatory Sequences, Ribonucleic Acid; D012091:Repetitive Sequences, Nucleic Acid; D000068103:Serine-Arginine Splicing Factors
D016633:Apolipoprotein A-II; D004268:DNA-Binding Proteins; D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H; D009687:Nuclear Proteins; D010750:Phosphoproteins; D016601:RNA-Binding Proteins; D038621:Regulatory Sequences, Ribonucleic Acid; D000068103:Serine-Arginine Splicing Factors