IFN-γ Mediates Enhancement of HIV Replication in Astrocytes by Inducing an Antagonist of the β-Catenin Pathway (DKK1) in a STAT 3-Dependent Manner This information is current as of September 24, 2021. Wei Li, Lisa J. Henderson, Eugene O. Major and Lena Al-Harthi J Immunol 2011; 186:6771-6778; Prepublished online 11 May 2011; doi: 10.4049/jimmunol.1100099 Downloaded from http://www.jimmunol.org/content/186/12/6771 References This article cites 41 articles, 10 of which you can access for free at: http://www.jimmunol.org/ http://www.jimmunol.org/content/186/12/6771.full#ref-list-1 Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists by guest on September 24, 2021 • Fast Publication! 4 weeks from acceptance to publication *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology IFN-g Mediates Enhancement of HIV Replication in Astrocytes by Inducing an Antagonist of the b-Catenin Pathway (DKK1) in a STAT 3-Dependent Manner Wei Li,*,1 Lisa J. Henderson,* Eugene O. Major,† and Lena Al-Harthi* Typically, IFN-g is an antiviral cytokine that inhibits the replication of many viruses, including HIV. However, in the CNS, IFN-g induces HIV-productive replication in astrocytes. Although astrocytes in vitro are refractory to HIV replication, recent in vivo evidence demonstrated that astrocytes are infected by HIV, and their degree of infection is correlated with proximity to activated macrophages/microglia. The ability of IFN-g to induce HIV replication in astrocytes suggests that the environmental milieu is critical in regulating the permissiveness of astrocytes to HIV infection. We evaluated the mechanism by which IFN-g relieves restricted HIV replication in astrocytes. We demonstrate that although astrocytes have robust endogenous b-catenin signaling, Downloaded from a pathway that is a potent inhibitor of HIV replication, IFN-g diminished b-catenin signaling in astrocytes by 40%, as evaluated by both active b-catenin protein expression and b-catenin-mediated T cell factor/lymphoid enhancer reporter (TOPflash) activity. Further, IFN-g–mediated inhibition of b-catenin signaling was dependent on its ability to induce an antagonist of the b-catenin signaling pathway, Dickkopf-related protein 1, in a STAT 3-dependent manner. Inhibition of STAT3 and Dickkopf-related protein 1 abrogated the ability of IFN-g to enhance HIV replication in astrocytes. These data demonstrated that IFN-g induces HIV replication in astrocytes by antagonizing the b-catenin pathway. To our knowledge, this is the first report to point to an intricate http://www.jimmunol.org/ cross-talk between IFN-g signaling and b-catenin signaling that may have biologic and virologic effects on HIV outcome in the CNS, as well as on broader processes where the two pathways interface. The Journal of Immunology, 2011, 186: 6771–6778. nterferon-g (IFN-g), a type II IFN, is a pleiotropic cytokine neuroinvasion and severity of neuropathogenesis in the human involved in antimicrobial and antitumor immunity by en- brain and the brain of SIV-infected macaques (4, 5). I hancing Ag presentation through MHC class I and class II, The majority of IFN-g effects are mediated by signaling through regulating a variety of genes, and facilitating proapoptotic re- the JAK–STAT pathway (6). IFN-g signaling through JAK–STAT sponses of infected cells (1). Although IFN-g is predominantly involves an initial step of IFN-g binding to its receptor, leading secreted by NK and NK T cells to activate macrophages and by to oligomerization of the IFN-g receptor subunits (IFNGR1 and by guest on September 24, 2021 effector CD4+ and CD8+ Ag-specific T cells, it is also secreted by IFNGR2), which causes phosphorylation and activation of JAKs. activated astrocytes and microglia in response to mechanical or JAK activation leads to phosphorylation and subsequent activa- ischemic injury (2). Further, IFN-g causes alteration in Ca2+ tion of STAT, which dimerize and translocate to the nucleus, waves in the astrocytic network, which is a marker of astrocyte where they bind g-activated sequences in the promoter of IFN-g– activation and may be important in the formation of synapses (3). regulated genes and, with cooperation from other transcriptional Although IFN-g is associated with enhanced anti-HIV immunity factors, such as breast cancer susceptibility gene 1 (BRCA1) and in the systemic compartment, in the CNS it is associated with HIV mini-chromosome maintenance protein 5 (MCM5), regulate IFN-g–responsive genes. Approximately 500 genes are regulated through the IFN-g–induced JAK–STAT pathway, including IFN- *Department of Immunology/Microbiology, Rush University Medical Center, Chi- inducible protein 10, GTPase, and suppressor of cytokine signal- cago, IL 60612; and †National Institute of Neurological Disease and Stroke, National ing I (1, 6). Seven STAT family members have been identified. Institutes of Health, Bethesda, MD 20892 STAT 3, in particular, is evident in reactive astrocytes and is linked 1 Current address: Department of Infectious Diseases, Beijing You’an Hospital, Cap- to neuroinflammatory responses in rodent models of ischemia ital Medical University, Beijing, China. and spinal cord injuries (7, 8). STAT 3 is activated by cytokines Received for publication January 14, 2011. Accepted for publication April 14, 2011. (IFN-g, IL-6, G-CSF) and growth hormones. It induces cell cycle This work was supported by Grants R01 NS060632 (to L.A.-H.) and F31 NS071999 progression, prevents apoptosis, and may be linked to oncogenesis (to L.J.H.) from the National Institutes of Health. It was also supported by the Chicago Developmental Center for AIDS Research (P30 AI 082151), a National through induction of proto-oncogenes, such as c-myc (9). Institutes of Health-funded program supported by the National Institute of Allergy HIV invades the brain early in the course of disease and leads and Infectious Diseases; the National Cancer Institute; the National Institute of Mental Health; the National Institute of Drug Abuse; the National Institute of Child to progressive neurologic impairments. Prior to the era of highly Health and Development; the National Heart, Lung, and Blood Institute; and the active antiretroviral therapy, HIV led to frank dementia/encepha- National Center for Complementary and Alternative Medicine. litis in ∼25% of HIV-infected individuals. Today, HIV causes Address correspondence and reprint requests to Dr. Lena Al-Harthi, Department of a milder, but much wider, spectrum of neurologic impairments, Immunology and Microbiology, Rush University Medical Center, 1735 West Harri- son Street, 614 Cohn, Chicago, IL 60612. E-mail address: [email protected] described as HIV-associated neurocognitive disorders (HAND). HAND symptoms include memory impairment, depression, Abbreviations used in this article: DN, dominant negative; FLUD, fludarabine; G3I, glycogen synthase kinase-3b inhibitor; GSK3b, glycogen synthase kinase-3b; HAD, tremors, psychosis, seizures, and behavioral changes, to name a HIV-associated dementia; HAND, HIV-associated neurocognitive disorders; HFA, few. Recent assessments from the CNS HIV Antiretroviral Ther- human fetal astrocyte; LEF, lymphoid enhancer; NIH, National Institutes of Health; PDA, progenitor-derived astrocyte; PFA, primal fetal astrocyte; S3I, STAT3 inhibitor; apy Effects Research (CHARTER) study (10) indicated that TCF, T cell factor. HAND occurs in 53% of HIV-infected individuals. HIV-mediated www.jimmunol.org/cgi/doi/10.4049/jimmunol.1100099 6772 INTERACTION BETWEEN IFN-g AND b-CATENIN SIGNALING IN CNS neuropathogenesis, depending on the severity of disease, includes Materials and Methods reactive astrocytosis, myelin pallor, and perturbations in synaptic Generation of primary human fetal astrocytes and human and dendritic density that may also include selective neuronal progenitor-derived astrocytes loss. The mechanism of HIV-mediated neurologic disorder is not Human fetal astrocytes (HFA), isolated at ∼20 wk gestation, were pur- entirely clear, but it is likely driven by both direct (active viral chased from Lonza (BioWhittaker, Walkersville, MD). Progenitor-derived replication) and indirect sequelae to HIV invasion of the brain. astrocytes (PDA) were generated from neural progenitor cells, as pre- Indirect mechanisms include dysregulation of glia, release of viral viously described (32). Briefly, progenitor cells were provided by Dr. proteins, and elevation of neurotoxic proteins (TNF-a, IL-6, IL- Eugene Major (National Institute of Neurological Disorders and Stroke, 1b, TGF-b, endothelin, glutamate) from resident brain cells and National Institutes of Health [NIH]) and seeded on poly-D-lysine–coated T-75 tissue culture flasks at 2 3 106 cells/flask. Cells were maintained infiltrating lymphocytes (11). in progenitor medium consisting of neurobasal media (Life Technolo- The primary targets of HIV infection in the CNS are infiltrating gies Invitrogen,