title: Rev inhibition strongly affects intracellular distribution of human immunodeficiency virus type 1 RNAs.
abstract: To define the human immunodeficiency virus type 1 (HIV-1) RNA maturation pathways, we analyzed the intracellular distribution of HIV-1 RNA and the viral regulatory proteins Rev and Tat in transfected COS cells and HIV-1-infected lymphoid C8166 cells by means of ultrastructural in situ hybridization using antisense RNA probes and immunoelectron microscopy. The intranuclear viral RNA occurs in ribonucleoprotein fibrils in the perichromatin and interchromatin regions. The simultaneous demonstration of Rev, Tat, Br-labeled RNA, and cellular proteins SC35 and CRM1 in such fibrils reveals the potential of Rev to associate with nascent HIV pre-mRNA and its splicing complex and transport machinery. In a rev-minus system, the env intron-containing, incompletely spliced viral RNAs are revealed only in the nucleus, indicating that Rev is required to initiate the transport to the cytoplasm. Moreover, env intron sequences frequently occur in the periphery of interchromatin granule clusters, while the probe containing the rev exon sequence does not associate with this nucleoplasmic domain. When cells were treated with the CRM1 inhibitor leptomycin B in the presence of Rev protein, the env intron containing HIV RNAs formed clusters throughout the nucleoplasm and accumulated at the nuclear pores. This suggests that Rev is necessary and probably also sufficient for the accumulation of incompletely spliced HIV RNAs at the nuclear pores while CRM1 is needed for translocation across the nuclear pore complex.
authors.affiliation: Centre of Electron Microscopy, University of Lausanne, 27 Bugnon, CH-1005 Lausanne, Switzerland.
authors.forename: Dusan
authors.initials: D
authors.lastname: Cmarko
authors.forename: Stig-Ove
authors.initials: SO
authors.lastname: Bøe
authors.forename: Catia
authors.initials: C
authors.lastname: Scassellati
authors.forename: Anne Marie
authors.initials: AM
authors.lastname: Szilvay
authors.forename: Svend
authors.initials: S
authors.lastname: Davanger
authors.forename: Xiang-Dong
authors.initials: XD
authors.lastname: Fu
authors.forename: Gunnar
authors.initials: G
authors.lastname: Haukenes
authors.forename: Karl-Henning
authors.initials: KH
authors.lastname: Kalland
authors.forename: Stanislav
authors.initials: S
authors.lastname: Fakan
mesh_terms: D021581:Active Transport, Cell Nucleus; D000818:Animals; D019556:COS Cells; D002467:Cell Nucleus / Q000378:metabolism; D002522:Chlorocebus aethiops; D005091:Exons; D015694:Gene Products, rev / Q000235:genetics / Q000378:metabolism*; D015696:Gene Products, tat / Q000235:genetics; D015752:Genes, env; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D007424:Intracellular Fluid / Q000378:metabolism; D007438:Introns; D028884:Karyopherins / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism; D010957:Plasmids; D012326:RNA Splicing; D012333:RNA, Messenger; D012367:RNA, Viral / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear; D012261:Ribonucleoproteins; D000068103:Serine-Arginine Splicing Factors; D014162:Transfection; D014771:Virion; D054321:rev Gene Products, Human Immunodeficiency Virus; D054322:tat Gene Products, Human Immunodeficiency Virus; D000097606:Exportin 1 Protein
chemical_list: D015694:Gene Products, rev; D015696:Gene Products, tat; D028884:Karyopherins; D009687:Nuclear Proteins; D012333:RNA, Messenger; D012367:RNA, Viral; D018160:Receptors, Cytoplasmic and Nuclear; D012261:Ribonucleoproteins; D000068103:Serine-Arginine Splicing Factors; D054321:rev Gene Products, Human Immunodeficiency Virus; D054322:tat Gene Products, Human Immunodeficiency Virus; D000097606:Exportin 1 Protein; C073288:SRSF2 protein, human