Effects of a Single Amino Acid Substitution within thep2 Region of Human Immunodeficiency Virus Type 1 on Packagingof Spliced ViralRNA

• DOI: 10.1128/jvi.77.24.12986-12995.2003
• Publication Date: 2003-11-25T22:59:20Z
• Authors: Rodney S. Russell; Ariel Roldan; Mervi Detorio; Jing Hu; Mark A. Wainberg; Chen Liang
• Keywords: 0301 basic medicine; Binding Sites; RNA Splicing; Virus Assembly; Gene Products, gag; Virus Replication; Cell Line; 3. Good health; Jurkat Cells; 03 medical and health sciences; Amino Acid Substitution; COS Cells; Chlorocebus aethiops; HIV-1; Animals; Humans; RNA, Viral; 5' Untranslated Regions; Dimerization; HeLa Cells

ABSTRACT Human immunodeficiency virus type 1 encapsidates two copies of viral genomic RNA in the form of a dimer. The dimerization process initiates via a 6-nucleotide palindrome that constitutes the loop of a viral RNA stem-loop structure (i.e., stem loop 1 [SL1], also termed the dimerization initiation site [DIS]) located within the 5′ untranslated region of the viral genome. We have now shown that deletion of the entire DIS sequence virtually eliminated viral replication but that this impairment was overcome by four second-site mutations located within the matrix (MA), capsid (CA), p2, and nucleocapsid (NC) regions of Gag. Interestingly, defective viral RNA dimerization caused by the ΔDIS deletion was not significantly corrected by these compensatory mutations, which did, however, allow the mutated viruses to package wild-type levels of this DIS-deleted viral RNA while excluding spliced viral RNA from encapsidation. Further studies demonstrated that the compensatory mutation T12I located within p2, termed MP2, sufficed to prevent spliced viral RNA from being packaged into the ΔDIS virus. Consistently, theΔ DIS-MP2 virus displayed significantly higher levels of infectiousness than did the ΔDIS virus. The importance of position T12 in p2 was further demonstrated by the identification of four point mutations,T12D, T12E, T12G, and T12P, that resulted in encapsidation of spliced viral RNA at significant levels. Taken together, our data demonstrate that selective packaging of viral genomic RNA is influenced by the MP2 mutation and that this represents a major mechanism for rescue of viruses containing the ΔDIS deletion.