RESUMO
Human T-cell leukemia virus type 1 (HTLV-1) is the etiologic agent of adult T-cell leukemia (ATL). The viral regulatory protein Tax1 plays a pivotal role in T-cell transformation and ATL development. Previous studies in our laboratory, using the yeast 2-hybrid approach to screen a T-cell library for Tax1-interacting partners, identified the cellular Four and a Half Lim domain protein 3 (FHL3) as a possible Tax1-interacting candidate. FHL3 is a member of the FHL family of proteins, which function as transcriptional coactivators and cytoskeleton regulators and have a role in cancer progression and development. The aim of this study was to investigate the physical and functional interaction between Tax1 and members of the FHL family of proteins. We show that Tax1 and FHL3 interact both in vitro and in vivo. Furthermore, both FHL1 and -2 also interact with Tax1. We have demonstrated that FHL3 enhances Tax1-mediated activation of the viral long terminal repeat (LTR) without affecting basal activity and that FHL1 to -3 regulate NF-κB activation by Tax1 in a cell-specific manner. In addition, we have found that the interaction between Tax1 and FHL1 to -3 affects the localization of these proteins, leading to their redistribution in cells. Tax1 also affected FHL3 cytoskeleton function by increasing FHL3-mediated cell spreading. Overall, our results suggest that the interaction between Tax1 and the FHL family alters both the transactivating activity and the subcellular localization of Tax1 and provide new insights into molecular mechanisms that underlie the oncogenic nature of this HTLV-1 protein.
Assuntos
Vírus Linfotrópico T Tipo 1 Humano/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas com Domínio LIM/metabolismo , Leucemia de Células T/virologia , Proteínas de Neoplasias/metabolismo , Animais , Western Blotting , Células COS , Chlorocebus aethiops , Imunoprecipitação da Cromatina , Citoesqueleto/metabolismo , Citoesqueleto/fisiologia , Imunofluorescência , Glutationa Transferase , Células HeLa , Humanos , Imunoprecipitação , Leucemia de Células T/metabolismo , Plasmídeos/genética , Técnicas do Sistema de Duplo-HíbridoRESUMO
BACKGROUND: In contrast with human T-cell leukemia virus type 1 (HTLV-1) that causes ATL (adult T-cell leukemia), HTLV-2 has not been causally linked to malignant disease. The minus strand of the HTLV genomes encode the regulatory proteins HTLV-1 bZIP factor (HBZ) for HTLV-1 and antisense protein of HTLV-2 (APH-2) for HTLV-2. Unlike the viral proteins Tax1 and Tax2, both HBZ and APH-2 are constitutively expressed in infected cells suggesting that they may play important roles in the pathogenesis of these viruses. To date, very little is known about the function of APH-2 except that it inhibits Tax2-mediated transcription of HTLV-2 genes. In the present study, we investigated the role of APH-2 in basal and Tax2B-mediated activation of the AP-1 pathway. RESULTS: We demonstrate that, unlike HBZ, APH-2 stimulates basal AP-1 transcription by interacting with c-Jun and JunB through its non-conventional bZIP domain. In addition, when Tax2 and APH-2 are co-expressed, they physically interact in vivo and in vitro and APH-2 acts as an inhibitor of Tax2-mediated activation of AP-1 transcription. CONCLUSIONS: This report is the first to document that HTLV-2 can modulate the AP-1 pathway. Altogether our results reveal that, in contrast with HBZ, APH-2 regulates AP-1 activity in a Tax2-dependant manner. As the AP-1 pathway is involved in numerous cellular functions susceptible to affect the life cycle of the virus, these distinct biological properties between HBZ and APH-2 may contribute to the differential pathogenic potential of HTLV-1 and HTLV-2.
Assuntos
Secretases da Proteína Precursora do Amiloide/metabolismo , Produtos do Gene tax/metabolismo , Vírus Linfotrópico T Tipo 2 Humano/metabolismo , Glicoproteínas de Membrana/metabolismo , Transdução de Sinais , Fator de Transcrição AP-1/metabolismo , Colagenases/genética , Regulação da Expressão Gênica , Humanos , Regiões Promotoras Genéticas , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Transporte Proteico , Proteínas Proto-Oncogênicas c-jun/genética , Proteínas Proto-Oncogênicas c-jun/metabolismo , Transcrição Gênica , Ativação TranscricionalRESUMO
SAMHD1 is a dNTP triphosphohydrolase (dNTPase) that impairs retroviral replication in a subset of non-cycling immune cells. Here we show that SAMHD1 is a redox-sensitive enzyme and identify three redox-active cysteines within the protein: C341, C350, and C522. The three cysteines reside near one another and the allosteric nucleotide binding site. Mutations C341S and C522S abolish the ability of SAMHD1 to restrict HIV replication, whereas the C350S mutant remains restriction competent. The C522S mutation makes the protein resistant to inhibition by hydrogen peroxide but has no effect on the tetramerization-dependent dNTPase activity of SAMHD1 in vitro or on the ability of SAMHD1 to deplete cellular dNTPs. Our results reveal that enzymatic activation of SAMHD1 via nucleotide-dependent tetramerization is not sufficient for the establishment of the antiviral state and that retroviral restriction depends on the ability of the protein to undergo redox transformations.