The multifaceted roles of PI(4,5)P2 on HIV-1 assembly

/ABSTRACT The HIV/AIDS pandemic has claimed more than 43 million lives worldwide. While effective antiretroviral therapies are widely available, poor adherence and the emergence of resistance remains a pressing challenge. Despite the notable advance in HIV-1 treatment over the decades, continuing basic research remains critically important to address the challenges posed by this rapidly evolving virus. HIV-1 assembly relies heavily on phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P2], a lipid that recruits the viral Gag protein to the plasma membrane (PM), where Gag orchestrates the assembly process. Intriguingly, our recent research suggests that PI(4,5)P2 has additional roles in HIV-1 assembly beyond simply recruiting Gag to the PM, indicating multiple, yet-undefined functions for this lipid in the viral life cycle. In the Aim 1 of this proposal, I will focus on elucidating the interplay between Gag multimerization and PI(4,5)P2 enrichment at HIV-1 assembly sites at the PM. I will also determine how this interplay facilitates the sorting of cellular proteins that either enhance or inhibit HIV-1 spread. Despite significant advances, the molecular mechanisms driving the incorporation of the HIV-1 glycoprotein Env into viral particles remain incompletely understood. In Aim 2, I will investigate the role played by PI(4,5)P2 in recruitment and incorporation of HIV-1 Env across various cell lines and primary CD4+ T cells. For this aim, I will use an innovative method for PI(4,5)P2 inhibition to probe this mechanism. Finally, our recent study raised the possibility that rather than promoting PI(4,5)P2 clustering, Gag can also be targeted to pre- existing PI(4,5)P2 pool at the PM. In Aim 3, during my independent research phase (R00), I will focus on elucidating whether Gag is addressed to pre-formed PI(4,5)P2 pools at the PM. In addition, I will elucidate the role of membrane contact sites (MCSs) in HIV-1 protein trafficking and assembly. MCSs, which facilitate direct communication between cellular organelles, have recently emerged as key players in non-canonical pathways for the targeted delivery of proteins and lipids. Interestingly, certain protein complexes that regulate ER-PM MCSs are known to enrich PI(4,5)P2, a lipid critical for HIV-1 assembly. Therefore, ER-PM MCSs may be a key mechanism for Gag trafficking to the PM, bypassing the canonical secretory pathway. By investigating these processes, my work will address significant gaps in our understanding of HIV-1 assembly that can push the field. Beyond research, I will engage in career development activities including technical training, leadership and diversity workshops, and regular advisory committee meetings to discuss progress and receive guidance to my transition to independence. Project Number: 1K99AI194970-01 | Fiscal Year: 2025 | NIH Institute/Center: National Institute of Allergy and Infectious Diseases (NIAID) | Principal Investigator: Ricardo de Souza Cardoso | Institution: UNIVERSITY OF MICHIGAN AT ANN ARBOR, ANN ARBOR, MI | Award Amount: $132,300 | Activity Code: K99 | Study Section: Acquired Immunodeficiency Syndrome Research Study Section[AIDS] View on NIH RePORTER: https://reporter.nih.gov/project-details/1K99AI19497001