title: A second exon splicing silencer within human immunodeficiency virus type 1 tat exon 2 represses splicing of Tat mRNA and binds protein hnRNP H.
abstract: An equilibrium between spliced and unspliced primary transcripts is essential for retrovirus multiplication. This equilibrium is maintained by the presence of inefficient splice sites. The A3 3'-splice site of human immunodeficiency virus type I (HIV-1) is required for Tat mRNA production. The infrequent utilization of this splice site has been attributed to the presence of a suboptimal polypyrimidine tract and an exonic splicing silencer (ESS2) in tat exon 2 approximately 60 nucleotides downstream of 3'-splice site A3. Here, using site-directed mutagenesis followed by analysis of splicing in vitro and in HeLa cells, we show that the 5' extremity of tat exon 2 contains a second exonic splicing silencer (ESS2p), which acts to repress splice site A3. The inhibitory property of this exonic silencer was active when inserted downstream of another HIV-1 3'-splice site (A2). Protein hnRNP H binds to this inhibitory element, and two U-to-C substitutions within the ESS2p element cause a decreased hnRNP H affinity with a concomitant increase in splicing efficiency at 3'-splice site A3. This suggests that hnRNP H is directly involved in splicing inhibition. We propose that hnRNP H binds to the HIV-1 ESS2p element and competes with U2AF(35) for binding to the exon sequence flanking 3'-splice site A3. This binding results in the inhibition of splicing at 3'-splice site A3.
authors.affiliation: Laboratoire de Maturation des Acide Ribo-Nucléotidique et Enzymologie Moléculaire, Unité Mixte de Recherche 7567 Université Henri Poincarré-CNRS, Boulevard des Aiguillettes, BP239, 54506 Vandoeuvre-lès-Nancy cedex, France.
authors.forename: S
authors.initials: S
authors.lastname: Jacquenet
authors.forename: A
authors.initials: A
authors.lastname: Méreau
authors.forename: P S
authors.initials: PS
authors.lastname: Bilodeau
authors.forename: L
authors.initials: L
authors.lastname: Damier
authors.forename: C M
authors.initials: CM
authors.lastname: Stoltzfus
authors.forename: C
authors.initials: C
authors.lastname: Branlant
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D002467:Cell Nucleus / Q000378:metabolism; D005091:Exons; D015696:Gene Products, tat / Q000235:genetics* / Q000378:metabolism; D015497:HIV-1 / Q000235:genetics*; D006367:HeLa Cells; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D012326:RNA Splicing; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012261:Ribonucleoproteins / Q000235:genetics* / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D054322:tat Gene Products, Human Immunodeficiency Virus
chemical_list: D015696:Gene Products, tat; D034481:Heterogeneous-Nuclear Ribonucleoprotein Group A-B; D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D012333:RNA, Messenger; D012261:Ribonucleoproteins; C439768:hnRNP A2; D054322:tat Gene Products, Human Immunodeficiency Virus