Structural elements in the Gag polyprotein of feline immunodeficiency virus involved in Gag self-association and assembly.
The Gag polyprotein of feline immunodeficiency virus (FIV) assembles at the plasma membrane of the infected cells. We aimed to identify the FIV Gag domains that interact and promote Gag multimerization. To do this we generated a series of Gag subdomains and tested their ability to associate with full-length Gag and be recruited into extracellular virus-like particles (VLPs). Removal of 37 residues from the C-terminus of FIV Gag and deletion of the N-terminal and central regions of the nucleocapsid (NC) domain attenuated but did not abrogate association with wild-type Gag, whereas a Gag mutant protein encompassing the matrix (MA) and capsid (CA) domains interacted poorly with full-length Gag. Association with wild-type Gag was abolished by deleting most of the NC together with the N-terminal 40 residues of the MA, which most likely reflects the inability of this Gag mutant to bind RNA. Notably, the CA-NC Gag subdomain both associated with wild-type Gag and was recruited into particles in a proportion close to 50 % of the total Gag-related protein mass of VLPs. Moreover, both a Gag protein lacking the C-terminal p2 peptide and a nonmyristoylated version of the polyprotein exhibited a transdominant-negative effect on the assembly of wild-type Gag. Analysis of Gag mutants carrying internal deletions within the CA revealed that the N-terminal and the C-terminal domains of the CA are necessary for Gag assembly. Our results demonstrate that the FIV CA-NC region constitutes the principal self-interaction domain of Gag and that the RNA-binding capacity of Gag is necessary for its multimerization.
Laboratorio de Virología, CONICET-Universidad de Belgrano (UB), Villanueva 1324, Buenos Aires C1426BMJ, Argentina.
Juan C
JC
Abdusetir Cerfoglio
Laboratorio de Virología, CONICET-Universidad de Belgrano (UB), Villanueva 1324, Buenos Aires C1426BMJ, Argentina.
Silvia A
SA
González
Laboratorio de Virología, CONICET-Universidad de Belgrano (UB), Villanueva 1324, Buenos Aires C1426BMJ, Argentina.
José L
JL
Affranchino
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D019556:COS Cells; D002213:Capsid / Q000378:metabolism; D036022:Capsid Proteins / Q000235:genetics; D002460:Cell Line; D002462:Cell Membrane / Q000821:virology; D002522:Chlorocebus aethiops; D015683:Gene Products, gag / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D016089:Immunodeficiency Virus, Feline / Q000235:genetics* / Q000472:pathogenicity; D008969:Molecular Sequence Data; D019251:Nucleocapsid / Q000235:genetics; D011485:Protein Binding / Q000235:genetics; D055503:Protein Multimerization / Q000235:genetics*; D017434:Protein Structure, Tertiary / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics; D051381:Rats; D016415:Sequence Alignment; D014616:Vaccinia virus / Q000235:genetics / Q000472:pathogenicity; D014763:Viral Matrix Proteins / Q000235:genetics; D019065:Virus Assembly / Q000235:genetics
D036022:Capsid Proteins; D015683:Gene Products, gag; D016601:RNA-Binding Proteins; D014763:Viral Matrix Proteins