Control of HIV-1 Infection By Ferroportin Q248H Mutation
Authors: Kumari Namita , Nouraie Seyed Mehdi , Lassiter Hatajai , Ahmad Asrar , Anastos Kathryn , Lazar Jason , Kassaye Seble , Aouizerat Bradley , French Audrey , Gange Stephen , Adimora Adaora , Weitzmann Neale , Fischl Margaret , Kempf Mirjam-Colette , Nekhai Sergei 
Published: February 2020

BACKGROUND: We recently showed that patients with Sickle Cell Disease (SCD), a hereditary hemolytic disorder, have low incidence of HIV-1 infection [1] and reduced ex vivo HIV-1 infection [2]. PBMC from SCD patients exhibited increased expression of iron export protein, ferroportin and reduced cellular iron levels leading to CDK2 inhibition, reduced SAMHD1 phosphorylation and increased expression of IkBα. Ferroportin expression is regulated by liver-produced hepcidin that facilitates ferroportin internalization and degradation. Ferroportin Q248H mutation has an allele frequency of 2.2-13.4% in African populations. We previously reported reduced sensitivity of ferroportin Q248H mutant to physiologic hepcidin concentrations in patients with sickle cell disease [3].
OBJECTIVES: To analyze the effect of ferroportin Q248H mutation on HIV-1 infection in vitro and in disease progression among a cohort of HIV-1 infected African-American women.
METHODS: HEK293 cells were used to express ferroportin Q248H mutant and test cellular ferritin and intracellular labile iron using calcein-AM. Confocal microscopy was used to visualize ferroportin expression. HIV-1 transcription was measured in 293T cells transfected with HIV-1 LTR-Luciferase vector and Tat expressing vector. Ex vivo infection was analyzed in monocyte-derived macrophages infected with VSVg-pseudotyped HIV-1 virus. Ferroportin Q248H mutation was genotyped using Thermo Fisher probe (C_25753769_10) and genotyping services at University of Utah.
RESULTS: We observed reduced intracellular iron in ferroportin Q248H expressing cells compared to WT ferroportin even when the cells were treated with hepcidin. In the absence of hepcidin, both WT ferroportin and Q248H ferroportin efficiently inhibited HIV-1 transcription and replication. Hepcidin induced HIV-1 transcription and replication in the cells with WT ferroportin but not Q248H mutant ferroportin. HIV-1 replication was reduced in primary macrophages obtained from patients with ferroportin Q248H mutation. To test whether expression of ferroportin Q248H offered protection from HIV-1 infection, we analyzed a cohort of HIV-1 infected women (WIHS). We genotyped 970 African-American subjects of whom 628 were HIV-1 infected and 342 were non-infected. The prevalence of Q248H hetero or homozygote mutations was 7.0% in non-infected and 11.8% among HIV-1 infected individuals (Odds Ratio=1.77, p=0.02). Analysis of HIV viral load showed significant lower viral load in the subjects with ferroportin Q248H mutation compared to WT.
CONCLUSIONS: Our findings point to the contribution of iron metabolism in HIV-1 restriction and the potential role of the ferroportin Q248H mutation in the regulation of HIV-1 infection in vivo.
ACKNOWLEDGMENTS: This work was supported by NIH Research Grants (1P50HL118006, 1R01HL125005, 5G12MD007597 and P30AI087714). We thank Women's Interagency HIV-1 study (WIHS) for sharing DNA samples and providing access to the clinical data. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
REFERENCES: Nouraie M, Nekhai S, Gordeuk VR. Sickle cell disease is associated with decreased HIV but higher HBV and HCV comorbidities in U.S. hospital discharge records: a cross-sectional study. Sex Transm Infect. 2012;88(7):528-533. Kumari N, Ammosova T, Diaz S, et al. Increased iron export by ferroportin induces restriction of HIV-1 infection in sickle cell disease. Blood Adv. 2016;1(3):170-183. Nekhai S, Xu M, Foster A, et al. Reduced sensitivity of the ferroportin Q248H mutant to physiological concentrations of hepcidin. Haematologica. 2013;98(3):455-463.

Disclosures
Anastos: NINR: Research Funding; NHGRI: Research Funding; NICHD: Research Funding; NIMH: Research Funding; NHLBI: Research Funding; NCI: Research Funding; NIAID: Research Funding; NINDS: Research Funding; NIDCR: Research Funding; NIMHD: Research Funding; NLM: Research Funding; Fogarty: Research Funding; NIDDK: Research Funding; NIA: Research Funding; NIAAA: Research Funding; NIDA: Research Funding.