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CD4 and MHC-I downregulation are conserved in primary HIV-1 Nef alleles from brain and lymphoid tissues, but Pak2 activation is highly variable.

ABSTRACT: HIV-1 compartmentalization in the CNS has been demonstrated for gag, pol, and env genes. However, little is known about tissue compartmentalization of nef genes and their functional characteristics in brain. We have cloned 97 nef genes and characterized 10 Nef proteins from autopsy brain and lymphoid tissues from 2 patients with AIDS and HIV-1-associated dementia. Distinct compartmentalization of brain versus lymphoid nef genes was demonstrated within each patient. CD4 and MHC-I downregulation were conserved in all tissue-derived Nefs. However, MHC-I downregulation by brain-derived Nefs was weaker than downregulation by lymphoid-derived Nefs. The motifs KEEE- or EKEE- at the PACS-1 binding site represented brain-specific signature patterns in these 2 patients and contributed to the reduced MHC-I downregulation activity of brain-derived Nefs from these patients. Pak2 association was highly variable in Nefs from both patients. Three of 10 tissue-derived Nefs coimmunoprecipitated activated Pak2, with strong association demonstrated for only 2 Nefs. The ability of Nef to associate with activated Pak2 did not correlate with brain or lymphoid tissue origin. Nef genes from viruses isolated from brain by coculture with PBMC were not closely related to sequences amplified directly from brain tissue, suggesting that viral selection or adaptation occurred during coculture. This study of tissue-derived HIV-1 Nefs demonstrates that CD4 and MHC-I downregulation are highly conserved Nef functions, while Pak2 association is variable in late stage AIDS patients.

SUBMITTER: Agopian K

PROVIDER: S-EPMC1995023 | biostudies-literature | 2007 Feb

REPOSITORIES: biostudies-literature

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CD4 and MHC-I downregulation are conserved in primary HIV-1 Nef alleles from brain and lymphoid tissues, but Pak2 activation is highly variable.

Agopian Kristin K Wei Bangdong L BL Garcia J Victor JV Gabuzda Dana D

Virology 20060918 1

HIV-1 compartmentalization in the CNS has been demonstrated for gag, pol, and env genes. However, little is known about tissue compartmentalization of nef genes and their functional characteristics in brain. We have cloned 97 nef genes and characterized 10 Nef proteins from autopsy brain and lymphoid tissues from 2 patients with AIDS and HIV-1-associated dementia. Distinct compartmentalization of brain versus lymphoid nef genes was demonstrated within each patient. CD4 and MHC-I downregulation w ...[more]

PMID: 16979207

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Project description:Patient-derived HIV-1 subtype B Nef clones downregulate HLA-A more efficiently than HLA-B. However, it remains unknown whether this property is common to Nef proteins across primate lentiviruses and how antiviral immune responses may be affected. We examined 263 Nef clones from diverse primate lentiviruses including different pandemic HIV-1 group M subtypes for their ability to downregulate major histocompatibility complex class A (MHC-A) and MHC-B from the cell surface. Though lentiviral Nef proteins differed markedly in their absolute MHC-A and MHC-B downregulation abilities, all lentiviral Nef lineages downregulated MHC-A, on average, 11 to 32% more efficiently than MHC-B. Nef genotype/phenotype analyses in a cohort of HIV-1 subtype C-infected patients (n = 168), together with site-directed mutagenesis, revealed Nef position 9 as a subtype-specific determinant of differential HLA-A versus HLA-B downregulation activity. Nef clones harboring nonconsensus variants at codon 9 downregulated HLA-B (though not HLA-A) significantly better than those harboring the consensus sequence at this site, resulting in reduced recognition of infected target cells by HIV-1-specific CD8+ effector cells in vitro Among persons expressing protective HLA class I alleles, carriage of Nef codon 9 variants was also associated with reduced ex vivo HIV-specific T cell responses. Our results demonstrate that Nef's inferior ability to downregulate MHC-B compared to that of MHC-A is conserved across primate lentiviruses and suggest that this property influences antiviral cellular immune responses.IMPORTANCE Primate lentiviruses encode the Nef protein that plays an essential role in establishing persistent infection in their respective host species. Nef interacts with the cytoplasmic region of MHC-A and MHC-B molecules and downregulates them from the infected cell surface to escape recognition by host cellular immunity. Using a panel of Nef alleles isolated from diverse primate lentiviruses including pandemic HIV-1 group M subtypes, we demonstrate that Nef proteins across all lentiviral lineages downregulate MHC-A approximately 20% more effectively than MHC-B. We further identify a naturally polymorphic site at Nef position 9 that contributes to the MHC-B downregulation function in HIV-1 subtype C and show that carriage of Nef variants with enhanced MHC-B downregulation ability is associated with reduced breadth and magnitude of MHC-B-restricted cellular immune responses in HIV-infected individuals. Our study underscores an evolutionarily conserved interaction between lentiviruses and primate immune systems that may contribute to pathogenesis.

Project description:UNLABELLED:Human immunodeficiency virus type 1 (HIV-1) Nef enhances virus replication and contributes to immune evasion in vivo, but the underlying molecular mechanisms remain incompletely defined. Nef interferes with host cell actin dynamics to restrict T lymphocyte responses to chemokine stimulation and T cell receptor engagement. This relies on the assembly of a labile multiprotein complex including the host kinase PAK2 that Nef usurps to phosphorylate and inactivate the actin-severing factor cofilin. Components of the exocyst complex (EXOC), an octameric protein complex involved in vesicular transport and actin remodeling, were recently reported to interact with Nef via the same molecular surface that mediates PAK2 association. Exploring the functional relevance of EXOC in Nef-PAK2 complex assembly/function, we found Nef-EXOC interactions to be specifically mediated by the PAK2 interface of Nef, to occur in infected human T lymphocytes, and to be conserved among lentiviral Nef proteins. In turn, EXOC was dispensable for direct downstream effector functions of Nef-associated PAK2. Surprisingly, PAK2 was essential for Nef-EXOC association, which required a functional Rac1/Cdc42 binding site but not the catalytic activity of PAK2. EXOC was dispensable for Nef functions in vesicular transport but critical for inhibition of actin remodeling and proximal signaling upon T cell receptor engagement. Thus, Nef exploits PAK2 in a stepwise mechanism in which its kinase activity cooperates with an adaptor function for EXOC to inhibit host cell actin dynamics. IMPORTANCE:Human immunodeficiency virus type 1 (HIV-1) Nef contributes to AIDS pathogenesis, but the underlying molecular mechanisms remain incompletely understood. An important aspect of Nef function is to facilitate virus replication by disrupting T lymphocyte actin dynamics in response to stimulation via its association with the host cell kinase PAK2. We report here that the molecular surface of Nef for PAK2 association also mediates interaction of Nef with EXOC and establish that PAK2 provides an essential adaptor function for the subsequent formation of Nef-EXOC complexes. PAK2 and EXOC specifically cooperate in the inhibition of actin dynamics and proximal signaling induced by T cell receptor engagement by Nef. These results establish EXOC as a functionally relevant Nef interaction partner, emphasize the suitability of the PAK2 interaction surface for future therapeutic interference with Nef function, and show that such strategies need to target activity-independent PAK2 functions.

Dataset Information

CD4 and MHC-I downregulation are conserved in primary HIV-1 Nef alleles from brain and lymphoid tissues, but Pak2 activation is highly variable.

Publications

CD4 and MHC-I downregulation are conserved in primary HIV-1 Nef alleles from brain and lymphoid tissues, but Pak2 activation is highly variable.

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