TY - JOUR
T1 - Influence of HIV-1 Genomic RNA on the Formation of Gag Biomolecular Condensates
AU - Monette, Anne
AU - Niu, Meijuan
AU - Maldonado, Rebecca Kaddis
AU - Chang, Jordan
AU - Lambert, Gregory S.
AU - Flanagan, John M.
AU - Cochrane, Alan
AU - Parent, Leslie J.
AU - Mouland, Andrew J.
N1 - Funding Information:
We thank lab members from all collaborating laboratories for comments on the work presented in this manuscript; Mathew Duguay and Christian Young of the LDI cell imaging and flow cytometry core facility for assistance with microscopy; and the NIH Reference and Reagent Program for antibodies and reagents. We thank Malgorzata Sudol, MS, for technical assistance. Research reported in this publication was supported by the National Institute On Drug Abuse of the National Institutes of Health under Award Number R21DA053689 to LJP with subaward to AJM. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. This work was also supported by grants from the Canadian Institutes of Health Research (CIHR) (FRN-162447 to A.J.M. and FRN- PJT-178165 to A.W.C.).
Publisher Copyright:
© 2023
PY - 2023/8/15
Y1 - 2023/8/15
N2 - Biomolecular condensates (BMCs) play an important role in the replication of a growing number of viruses, but many important mechanistic details remain to be elucidated. Previously, we demonstrated that the pan-retroviral nucleocapsid (NC) and HIV-1 pr55Gag (Gag) proteins phase separate into condensates, and that HIV-1 protease (PR)-mediated maturation of Gag and Gag-Pol precursor proteins yields self-assembling BMCs that have HIV-1 core architecture. Using biochemical and imaging techniques, we aimed to further characterize the phase separation of HIV-1 Gag by determining which of its intrinsically disordered regions (IDRs) influence the formation of BMCs, and how the HIV-1 viral genomic RNA (gRNA) could influence BMC abundance and size. We found that mutations in the Gag matrix (MA) domain or the NC zinc finger motifs altered condensate number and size in a salt-dependent manner. Gag BMCs were also bimodally influenced by the gRNA, with a condensate-promoting regime at lower protein concentrations and a gel dissolution at higher protein concentrations. Interestingly, incubation of Gag with CD4+ T cell nuclear lysates led to the formation of larger BMCs compared to much smaller ones observed in the presence of cytoplasmic lysates. These findings suggest that the composition and properties of Gag-containing BMCs may be altered by differential association of host factors in nuclear and cytosolic compartments during virus assembly. This study significantly advances our understanding of HIV-1 Gag BMC formation and provides a foundation for future therapeutic targeting of virion assembly.
AB - Biomolecular condensates (BMCs) play an important role in the replication of a growing number of viruses, but many important mechanistic details remain to be elucidated. Previously, we demonstrated that the pan-retroviral nucleocapsid (NC) and HIV-1 pr55Gag (Gag) proteins phase separate into condensates, and that HIV-1 protease (PR)-mediated maturation of Gag and Gag-Pol precursor proteins yields self-assembling BMCs that have HIV-1 core architecture. Using biochemical and imaging techniques, we aimed to further characterize the phase separation of HIV-1 Gag by determining which of its intrinsically disordered regions (IDRs) influence the formation of BMCs, and how the HIV-1 viral genomic RNA (gRNA) could influence BMC abundance and size. We found that mutations in the Gag matrix (MA) domain or the NC zinc finger motifs altered condensate number and size in a salt-dependent manner. Gag BMCs were also bimodally influenced by the gRNA, with a condensate-promoting regime at lower protein concentrations and a gel dissolution at higher protein concentrations. Interestingly, incubation of Gag with CD4+ T cell nuclear lysates led to the formation of larger BMCs compared to much smaller ones observed in the presence of cytoplasmic lysates. These findings suggest that the composition and properties of Gag-containing BMCs may be altered by differential association of host factors in nuclear and cytosolic compartments during virus assembly. This study significantly advances our understanding of HIV-1 Gag BMC formation and provides a foundation for future therapeutic targeting of virion assembly.
UR - http://www.scopus.com/inward/record.url?scp=85165385117&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85165385117&partnerID=8YFLogxK
U2 - 10.1016/j.jmb.2023.168190
DO - 10.1016/j.jmb.2023.168190
M3 - Article
C2 - 37385580
AN - SCOPUS:85165385117
SN - 0022-2836
VL - 435
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
IS - 16
M1 - 168190
ER -