title: Discovery and characterization of auxiliary proteins encoded by type 3 simian T-cell lymphotropic viruses.
abstract: Human T-cell lymphotropic virus type 1 (HTLV-1) and HTLV-2 encode auxiliary proteins that play important roles in viral replication, viral latency, and immune escape. The presence of auxiliary protein-encoding open reading frames (ORFs) in HTLV-3, the latest HTLV to be discovered, is unknown. Simian T-cell lymphotropic virus type 3 (STLV-3) is almost identical to HTLV-3. Given the lack of HTLV-3-infected cell lines, we took advantage of STLV-3-infected cells and of an STLV-3 molecular clone to search for the presence of auxiliary transcripts. Using reverse transcriptase PCR (RT-PCR), we first uncovered the presence of three unknown viral mRNAs encoding putative proteins of 5, 8, and 9 kDa and confirmed the presence of the previously reported RorfII transcript. The existence of these viral mRNAs was confirmed by using splice site-specific RT-PCR with ex vivo samples. We showed that p5 is distributed throughout the cell and does not colocalize with a specific organelle. The p9 localization is similar to that of HTLV-1 p12 and induced a strong decrease in the calreticulin signal, similarly to HTLV-1 p12. Although p8, RorfII, and Rex-3 share an N-terminal sequence that is predicted to contain a nucleolar localization signal (NoLS), only p8 is found in the nucleolus. The p8 location in the nucleolus is linked to a bipartite NoLS. p8 and, to a lesser extent, p9 repressed viral expression but did not alter Rex-3-dependent mRNA export. Using a transformation assay, we finally showed that none of the STLV-3 auxiliary proteins had the ability to induce colony formation, while both Tax-3 and antisense protein of HTLV-3 (APH-3) promoted cellular transformation. Altogether, these results complete the characterization of the newly described primate T-lymphotropic virus type 3 (PTLV-3).
  
  
  OBJECTIVE
  Together with their simian counterparts, HTLVs form the primate T-lymphotropic viruses. HTLVs arose from interspecies transmission between nonhuman primates and humans. HTLV-1 and HTLV-2 encode auxiliary proteins that play important roles in viral replication, viral latency, and immune escape. The presence of ORFs encoding auxiliary proteins in HTLV-3 or STLV-3 genomes was unknown. Using in silico analyses, ex vivo samples, or in vitro experiments, we have uncovered the presence of 3 previously unknown viral mRNAs encoding putative proteins and confirmed the presence of a previously reported viral transcript. We characterized the intracellular localization of the four proteins. We showed that two of these proteins repress viral expression but that none of them have the ability to induce colony formation. However, both Tax and the antisense protein APH-3 promote cell transformation. Our results allowed us to characterize 4 new retroviral proteins for the first time.
authors.affiliation: Equipe Oncogenèse Rétrovirale, Lyon, France Equipe Labellisée Ligue Nationale Contre le Cancer, Lyon, France International Center for Research in Infectiology, INSERM U1111-CNRS UMR5308, Lyon, France Ecole Normale Supérieure de Lyon, Lyon, France Université Lyon 1, Lyon, France.
authors.forename: Jocelyn
authors.initials: J
authors.lastname: Turpin
authors.affiliation: Equipe Oncogenèse Rétrovirale, Lyon, France Equipe Labellisée Ligue Nationale Contre le Cancer, Lyon, France International Center for Research in Infectiology, INSERM U1111-CNRS UMR5308, Lyon, France Ecole Normale Supérieure de Lyon, Lyon, France Université Lyon 1, Lyon, France.
authors.forename: Chloé
authors.initials: C
authors.lastname: Journo
authors.affiliation: Epidémiologie et Physiopathologie des Virus Oncogènes, CNRS UMR 3569, Pasteur Institute, Paris, France.
authors.forename: Nga Ling
authors.initials: NL
authors.lastname: Ko
authors.affiliation: Equipe Oncogenèse Rétrovirale, Lyon, France Equipe Labellisée Ligue Nationale Contre le Cancer, Lyon, France International Center for Research in Infectiology, INSERM U1111-CNRS UMR5308, Lyon, France Ecole Normale Supérieure de Lyon, Lyon, France Université Lyon 1, Lyon, France Master Biosciences ENS Lyon, Lyon, France.
authors.forename: Flore
authors.initials: F
authors.lastname: Sinet
authors.affiliation: Molecular and Cellular Epigenetics (GIGA) and Molecular Biology (Gembloux Agro-Bio Tech), University of Liège, Liège, Belgium.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Carpentier
authors.affiliation: Equipe Oncogenèse Rétrovirale, Lyon, France Equipe Labellisée Ligue Nationale Contre le Cancer, Lyon, France International Center for Research in Infectiology, INSERM U1111-CNRS UMR5308, Lyon, France Ecole Normale Supérieure de Lyon, Lyon, France Université Lyon 1, Lyon, France Master Biosciences ENS Lyon, Lyon, France.
authors.forename: Amandine
authors.initials: A
authors.lastname: Galioot
authors.affiliation: Virus Tumor Biology Section, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, Maryland, USA.
authors.forename: Dustin
authors.initials: D
authors.lastname: Edwards
authors.affiliation: University of Leuven, Department of Microbiology and Immunology, Rega Institute for Medical Research, Clinical and Epidemiological Virology, Leuven, Belgium, and Centro de Malária e outras Doenças Tropicais and Unidade de Microbiologia, Instituto de Higiene e Medicina Tropical, Universidade Nova de Lisboa, Lisbon, Portugal.
authors.forename: Anne-Mieke
authors.initials: AM
authors.lastname: Vandamme
authors.affiliation: Ecole Normale Supérieure de Lyon, Lyon, France Laboratoire de Biologie Moléculaire de la Cellule, Unité Mixte de Recherche 5239, Centre National de la Recherche Scientifique, UMS3444 Biosciences Lyon-Gerland, Lyon, France.
authors.forename: Louis
authors.initials: L
authors.lastname: Gazzolo
authors.affiliation: Ecole Normale Supérieure de Lyon, Lyon, France Laboratoire de Biologie Moléculaire de la Cellule, Unité Mixte de Recherche 5239, Centre National de la Recherche Scientifique, UMS3444 Biosciences Lyon-Gerland, Lyon, France.
authors.forename: Madeleine
authors.initials: M
authors.lastname: Duc Dodon
authors.affiliation: Epidémiologie et Physiopathologie des Virus Oncogènes, CNRS UMR 3569, Pasteur Institute, Paris, France.
authors.forename: Antoine
authors.initials: A
authors.lastname: Gessain
authors.affiliation: National Center for Biodefense and Infectious Diseases, George Mason University, Manassas, Virginia, USA.
authors.forename: Fatah
authors.initials: F
authors.lastname: Kashanchi
authors.affiliation: UMS 3537 CNRS-AMU, Marseille, France.
authors.forename: Ivan
authors.initials: I
authors.lastname: Balansard
authors.affiliation: Station de Primatologie-UPS846-CNRS, Rousset sur Arc, France.
authors.forename: Romain
authors.initials: R
authors.lastname: Lacoste
authors.affiliation: Equipe Oncogenèse Rétrovirale, Lyon, France Equipe Labellisée Ligue Nationale Contre le Cancer, Lyon, France International Center for Research in Infectiology, INSERM U1111-CNRS UMR5308, Lyon, France Ecole Normale Supérieure de Lyon, Lyon, France Université Lyon 1, Lyon, France renaud.mahieux@ens-lyon.fr.
authors.forename: Renaud
authors.initials: R
authors.lastname: Mahieux
mesh_terms: D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000737:chemistry; D003600:Cytosol / Q000737:chemistry; D020869:Gene Expression Profiling; D006801:Humans; D008970:Molecular Weight; D020133:Reverse Transcriptase Polymerase Chain Reaction; D056185:Simian T-lymphotropic virus 3 / Q000235:genetics* / Q000502:physiology*; D014764:Viral Proteins / Q000032:analysis* / Q000737:chemistry / Q000235:genetics*
chemical_list: D014764:Viral Proteins