| Autor: |
Medica S; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA.; Department of Molecular Microbiology and Immunology, Oregon Health and Science University, Portland, Oregon, USA., Denton M; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Diggins NL; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Kramer-Hansen O; Department of Biomedical Sciences Molecular Pharmacology, University of Copenhagen, Copenhagen, Denmark., Crawford LB; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Mayo AT; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Perez WD; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Daily MA; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Parkins CJ; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Slind LE; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Pung LJ; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Weber WC; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA.; Department of Molecular Microbiology and Immunology, Oregon Health and Science University, Portland, Oregon, USA., Jaeger HK; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Streblow ZJ; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Sulgey G; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Kreklywich CN; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Alexander T; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Rosenkilde MM; Department of Biomedical Sciences Molecular Pharmacology, University of Copenhagen, Copenhagen, Denmark., Caposio P; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Hancock MH; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA., Streblow DN; Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, Oregon, USA.; Department of Molecular Microbiology and Immunology, Oregon Health and Science University, Portland, Oregon, USA.; Division of Pathobiology and Immunology, Oregon National Primate Research Center, Beaverton, Oregon, USA. |
| Abstrakt: |
The human cytomegalovirus (HCMV) encoded chemokine receptor US28 plays a critical role in viral pathogenesis, mediating several processes such as cellular migration, differentiation, transformation, and viral latency and reactivation. Despite significant research examining the signal transduction pathways utilized by US28, the precise mechanism by which US28 activates these pathways remains unclear. We performed a mutational analysis of US28 to identify signaling domains that are critical for functional activities. Our results indicate that specific residues within the third intracellular loop (ICL3) of US28 are major determinants of G-protein coupling and downstream signaling activity. Alanine substitutions at positions S218, K223, and R225 attenuated US28-mediated activation of MAPK and RhoA signal transduction pathways. Furthermore, we show that mutations at positions S218, K223, or R225 result in impaired coupling to multiple Gα isoforms. However, these substitutions did not affect US28 plasma membrane localization or the receptor internalization rate. Utilizing CD34 + HPC models, we demonstrate that attenuation of US28 signaling via mutation of residues within the ICL3 region results in an inability of the virus to efficiently reactivate from latency. These results were recapitulated in vivo , utilizing a humanized mouse model of HCMV infection. Together, our results provide new insights into the mechanism by which US28 manipulates host signaling networks to mediate viral latency and reactivation. The results reported here will guide the development of targeted therapies to prevent HCMV-associated disease.IMPORTANCEHuman cytomegalovirus (HCMV) is a β-herpesvirus that infects between 44% and 100% of the world population. Primary infection is typically asymptomatic and results in the establishment of latent infection within CD34 + hematopoietic progenitor cells (HPCs). However, reactivation from latent infection remains a significant cause of morbidity and mortality in immunocompromised individuals. The viral chemokine receptor US28 influences various cellular processes crucial for viral latency and reactivation, yet the precise mechanism by which US28 functions remains unclear. Through mutational analysis, we identified key residues within the third intracellular loop (ICL3) of US28 that govern G-protein coupling, downstream signaling, and viral reactivation in vitro and in vivo . These findings offer novel insights into how US28 manipulates host signaling networks to regulate HCMV latency and reactivation and expand our understanding of HCMV pathogenesis. |
