Contribution of the Gag-Pol transframe domain p6* and its coding sequence to morphogenesis and replication of human immunodeficiency virus type 1.
The human immunodeficiency virus type-1 (HIV-1) transframe domain p6* is located between the nucleocapsid protein (NC) and the protease (PR) within the Gag-Pol precursor. This flexible, 68-amino-acid HIV-1 p6* domain has been suggested to negatively interfere with HIV PR activity in vitro proposing a contribution of either the C-terminal p6* tetrapeptide, internal cryptic PR cleavage sites, or a zymogen-related mechanism to a regulated PR activation. To assess these hypotheses in the viral context, a series of recombinant HX10-based provirus constructs has been established with clustered amino acid substitutions throughout the entire p6* coding sequence. Comparative analysis of the mutant proviral clones in different cell culture systems revealed that mutations within the well-conserved amino-terminal p6* region modified the Gag/Gag-Pol ratio and thus resulted in the release of viruses with impaired infectivity. Clustered amino acid substitutions destroying (i) the predicted cryptic PR cleavage sites or (ii) homologies to the pepsinogen propeptide did not influence viral replication in cell culture, whereas substitutions of the carboxyl-terminal p6* residues 62 to 68 altering proper release of the mature PR from the Gag-Pol precursor drastically reduced viral infectivity. Thus, the critical contribution of p6* and overlapping cis-acting sequence elements to timely regulated virus maturation and infectivity is closely linked to precise ribosomal frameshifting and proper N-terminal release of the viral PR from the Gag-Pol precursor, clearly disproving the hypothesis that sequence motifs in the central part of p6* modulate PR activation and viral infectivity.
Paulus
Christina
C
Institute of Medical Microbiology and Hygiene, D-93053 Regensburg, Germany.
Ludwig
Christine
C
Wagner
Ralf
R
D000595:Amino Acid Sequence; D001483:Base Sequence; D004789:Enzyme Activation; D016368:Frameshift Mutation; D015685:Fusion Proteins, gag-pol / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D016333:HIV Protease / Q000378:metabolism; D015497:HIV-1 / Q000254:growth & development / Q000472:pathogenicity / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D009024:Morphogenesis; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D014779:Virus Replication / Q000502:physiology*
D015685:Fusion Proteins, gag-pol; D010446:Peptide Fragments; D016333:HIV Protease