ABSTRACT
HIN-200 proteins are interferon-inducible proteins capable of regulating cell growth, senescence, differentiation and death. Using a combination of in silico analysis of NCBI EST databases and screening of murine C57BL/6 cDNA libraries we isolated novel murine HIN-200 cDNAs designated Ifi206S and Ifi206L encoding two putative mRNA splice variants. The p206S and p206L protein isoforms have a modular domain structure consisting of an N-terminal PAAD/DAPIN/Pyrin domain, a region rich in serine, threonine and proline residues and a C-terminal 200 B domain characteristic of other HIN-200 proteins. Ifi206 mRNA was detected only in the spleen and lung of BALB/c and C57BL/6 mice and expression was up-regulated by both types I and II IFN subtypes. p206 protein was predominantly expressed in the cytoplasm and addition of LMB, a CRM1 dependent nuclear export inhibitor, caused p206 to accumulate in the nucleus. Unlike other human and mouse HIN-200 proteins that contain only a single 200 amino acid domain, overexpression of p206 impaired the clonogenic growth of tumour cell lines. Thus, p206 represents the newest HIN-200 family member discovered. It has distinct and restricted pattern of expression however maintains many of the hallmarks of HIN-200 proteins including the presence of a characteristic 200 X domain, induction by interferon and an ability to suppress tumour cell growth.
Subject(s)
Nuclear Proteins/metabolism , Tumor Suppressor Proteins/metabolism , 3T3 Cells , Alternative Splicing , Animals , Cell Line, Tumor , Cell Proliferation , Cloning, Molecular , Cytoplasm/metabolism , Gene Library , Interferon Type I/pharmacology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Nuclear Proteins/genetics , Organ Specificity , Protein Conformation , RNA, Messenger/metabolism , Recombinant Proteins , Transfection , Tumor Suppressor Proteins/geneticsABSTRACT
Astrocytes are the most numerous cell type in the brain, and their physiological roles are essential for normal brain function. Studies of post-mortem brain tissue samples from individuals with AIDS have revealed that a small proportion of astrocytes are infected by HIV-1 which is linked to the development of HIV-associated dementia (HIVD), a frequent clinical manifestation of HIV-1 disease affecting up to 20% of infected adults. However, astrocyte infection by HIV-1 in vivo is generally non-productive, and can only be readily detected by sensitive techniques that detect HIV-1 RNA or proviral DNA. Similarly, primary astrocyte cultures and astrocytic cell lines can be permissive to infection by HIV-1 strains, but are refractory to efficient HIV-1 expression. In efforts to delineate the molecular mechanisms underlying the "restricted" infection, several studies have demonstrated that efficient HIV-1 replication is blocked in astrocytes at different steps of the virus life cycle, including virus entry, reverse transcription, nucleocytoplasmic HIV-1 RNA transport, translation of viral RNA, and maturation of progeny virions. However, the relative importance of each of these possible replication blocks in restricting HIV-1 replication in astrocytes is unclear. Moreover, how restricted astrocyte infection contributes to the development of HIVD is unknown. This review surveys the current in vitro models of restricted HIV-1 replication in astrocytes, and provides an analysis of the available evidence supporting a role for astrocyte infection in the pathogenesis of HIVD. A greater understanding of the fate of HIV-1 in astrocytes may assist in the identification of viral reservoirs in the central nervous system, novel therapies for the treatment of HIVD, and also novel strategies to suppress HIV-1 replication in CD4+ cells of the immune system.
Subject(s)
AIDS Dementia Complex/virology , Astrocytes/virology , HIV-1/physiology , Cytokines , Gene Products, rev/physiology , HIV Long Terminal Repeat/physiology , Humans , Neurotoxins , Protein Biosynthesis , RNA-Binding Proteins/physiology , Receptors, HIV/physiology , Transcription, Genetic , Virus Replication , eIF-2 Kinase/metabolism , rev Gene Products, Human Immunodeficiency VirusABSTRACT
Acute human immunodeficiency virus type 1 (HIV-1) replication in astrocytes produces minimal new virus particles due, in part, to inefficient translation of viral structural proteins despite high levels of cytoplasmic viral mRNA. We found that a highly reactive double-stranded (ds) RNA-binding protein kinase (PKR) response in astrocytes underlies this inefficient translation of HIV-1 mRNA. The dsRNA elements made during acute replication of HIV-1 in astrocytes triggers PKR activation and the specific inhibition of HIV-1 protein translation. The heightened PKR response results from relatively low levels of the cellular antagonist of PKR, the TAR RNA binding protein (TRBP). Efficient HIV-1 production was restored in astrocytes by inhibiting the innate PKR response to HIV-1 dsRNA with dominant negative PKR mutants, or PKR knockdown by siRNA gene silencing. Increasing the expression of TRBP in astrocytes restored acute virus production to levels comparable to those observed in permissive cells. Therefore, the robust innate PKR antiviral response in astrocytes results from relatively low levels of TRBP expression and contributes to their restricted infection. Our findings highlight TRBP as a novel cellular target for therapeutic interventions to block productive HIV-1 replication in cells that are fully permissive for HIV-1 infection.