A Stretch of Unpaired Purines in the Leader Region of Simian Immunodeficiency Virus (SIV) Genomic RNA is Critical for its Packaging into Virions

• Published in: Journal of molecular biology Vol. 433; no. 23; p. 167293
• Main Authors: Pillai, Vineeta N., Ali, Lizna Mohamed, Prabhu, Suresha G., Krishnan, Anjana, Chameettachal, Akhil, Pitchai, Fathima Nuzra Nagoor, Mustafa, Farah, Rizvi, Tahir A.
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 19.11.2021
• Subjects: Animals; Base Composition; Base Pairing; Gene Expression Regulation, Viral; Genome, Viral - genetics; Nucleic Acid Conformation; Purines; Retroviruses; RNA packaging; RNA secondary structure; RNA, Viral - chemistry; RNA, Viral - genetics; Simian Acquired Immunodeficiency Syndrome - virology; Simian Immunodeficiency Virus (SIV); Simian Immunodeficiency Virus - physiology; Single-stranded purines (ssPurines); Virus Assembly; Virus Replication; Single-stranded purines (ssPurines); RNA packaging; Retroviruses; RNA secondary structure; Simian Immunodeficiency Virus (SIV)
• ISSN: 0022-2836; 1089-8638
• DOI: 10.1016/j.jmb.2021.167293

[Display omitted] •Single stranded purines (ssPurines) are crucial for retroviral gRNA packaging.•ssPurines I-IV are identified in the SHAPE-validated structure of SIV 5′ end gRNA.•ssPurines I, III, & IV may facilitate but are not essential for SIV gRNA packaging.•ssPurine II and its unpaired conformation is indispensable for SIV gRNA packaging.•ssPurine II may facilitate SIV gRNA packaging by acting as a Gag binding site. Simian immunodeficiency virus (SIV) is an important lentivirus used as a non-human primate model to study HIV replication, and pathogenesis of human AIDS, as well as a potential vector for human gene therapy. This study investigated the role of single-stranded purines (ssPurines) as potential genomic RNA (gRNA) packaging determinants in SIV replication. Similar ssPurines have been implicated as important motifs for gRNA packaging in many retroviruses like, HIV-1, MPMV, and MMTV by serving as Gag binding sites during virion assembly. In examining the secondary structure of the SIV 5′ leader region, as recently deduced using SHAPE methodology, we identified four specific stretches of ssPurines (I-IV) in the region that harbors major packaging determinants of SIV. The significance of these ssPurine motifs were investigated by mutational analysis coupled with a biologically relevant single round of replication assay. These analyses revealed that while ssPurine II was essential, the others (ssPurines I, III, & IV) did not significantly contribute to SIV gRNA packaging. Any mutation in the ssPurine II, such as its deletion or substitution, or other mutations that caused base pairing of ssPurine II loop resulted in near abrogation of RNA packaging, further substantiating the crucial role of ssPurine II and its looped conformation in SIV gRNA packaging. Structure prediction analysis of these mutants further corroborated the biological results and further revealed that the unpaired nature of ssPurine II is critical for its function during SIV RNA packaging perhaps by enabling it to function as a specific binding site for SIV Gag.