title: Molecular and cellular correlates of the CIITA-mediated inhibition of HTLV-2 Tax-2 transactivator function resulting in loss of viral replication.
abstract: BACKGROUND
  MHC class II transactivator CIITA inhibits the function of HTLV-2 Tax-2 viral transactivator and, consequently, the replication of the virus in infected cells. Moreover overexpression of the nuclear factor NF-YB, that cooperates with CIITA for the expression of MHC class II genes, results also in inhibition of Tax-2 transactivation. The purpose of this investigation was to assess the cellular and molecular basis of the CIITA-mediated inhibition on Tax-2, and the relative role of NF-YB in this phenomenon.
  
  
  METHODS
  By co-immunoprecipitation of lysates from 293T cells cotransfected with CIITA or fragments of it, and Tax-2 it was assessed whether the two factors interact in vivo. A similar approach was used to assess Tax-2-NF-YB interaction. In parallel, deletion fragments of CIITA were tested for the inhibition of Tax-2-dependent HTLV-2 LTR-luciferase transactivation. Subcellular localization of CIITA and Tax-2 was investigated by immunofluorescence and confocal microscopy.
  
  
  RESULTS
  CIITA and Tax-2 interact in vivo through at least two independent regions, at the 1-252 N-term and at the 410-1130 C-term, respectively. Interestingly only the 1-252 N-term region mediates Tax-2 functional inhibition. CIITA and Tax-2 are localized both in the cytoplasm and in the nucleus, when separately expressed. Instead, when coexpressed, most of Tax-2 colocalize with CIITA in cytoplasm and around the nuclear membrane. The Tax-2 minor remaining nuclear portion also co-localizes with CIITA. Interestingly, when CIITA nucleus-cytoplasm shuttling is blocked by leptomycin B treatment, most of the Tax-2 molecules are also blocked and co-localize with CIITA in the nucleus, suggesting that CIITA-Tax-2 binding does not preclude Tax-2 entry into the nucleus.Finally, the nuclear factor NF-YB, also strongly binds to Tax-2. Notably, although endogenous NF-YB does not inhibit Tax-2-dependent HTLV-2 LTR transactivation, it still binds to Tax-2, and in presence of CIITA, this binding seems to increase.
  
  
  CONCLUSIONS
  These results strongly suggest that CIITA inhibit Tax-2 by binding the viral transactivator both directly or through a tripartite interaction with NF-YB in. CIITA is therefore a viral restriction factor for HTLV-2 and this open the possibility to control HTLV-2 viral replication and spreading by the controlled induction of CIITA in infected cells.
authors: Department of Experimental Medicine, University of Insubria, Varese, Italy.
authors: Chiara
authors: C
authors: Orlandi
authors: Greta
authors: G
authors: Forlani
authors: Giovanna
authors: G
authors: Tosi
authors: Roberto S
authors: RS
authors: Accolla
mesh_terms: D023081:CCAAT-Binding Factor / Q000378:metabolism; D016356:Gene Products, tax / Q000037:antagonists & inhibitors* / Q000378:metabolism; D057809:HEK293 Cells; D015367:Human T-lymphotropic virus 2 / Q000502:physiology*; D006801:Humans; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D021381:Protein Transport; D013347:Subcellular Fractions / Q000378:metabolism; D015534:Trans-Activators / Q000737:chemistry / Q000378:metabolism*; D014779:Virus Replication / Q000502:physiology*
chemical_list: D023081:CCAAT-Binding Factor; D016356:Gene Products, tax; C083728:MHC class II transactivator protein; D009687:Nuclear Proteins; D015534:Trans-Activators; C514971:tax protein, Human T-lymphotrophic virus 2