RESUMO
In our search for new partners of the HIV-1 envelope glycoprotein (Env), we found that the cytoplasmic domain of the TMgp41 (TMgp41 CD) subunit of HIV-1 Env interacted with Luman, a transcription factor of the CREB/ATF family. Luman is anchored in the endoplasmic reticulum membrane and subjected to activation by regulated intramembrane proteolysis (RIP). The RIP process permits the release of the activated amino-terminal fragment of Luman into the cytoplasm, and its import into the nucleus. Here, we demonstrate that interaction between the TMgp41 CD and Luman requires a region encompassing the b-Zip and TM domains of Luman and decreases the stability of this factor. Moreover, we found that overexpression of a constitutively active form of Luman in cells transfected with HXB2R HIV-1 provirus decreased the intracellular expression of Gag and Env and led to a decrease in virion release. This negative effect of activated Luman on HIV-1 production was correlated to the inhibition of Tat transactivation of the HIV-1 LTR, which might be related to an interaction of activated Luman with Tat. Altogether, these results show that Luman acts as a partner of two major HIV-1 proteins: the TMgp41 Env subunit and Tat. The interaction between the TMgp41 subunit of Env and Luman affects the stability of the full-length Luman protein, the precursor of the activated, nuclear form of Luman, which acts negatively on Tat-mediated HIV-1 transactivation.
Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Produtos do Gene tat/metabolismo , Proteína gp41 do Envelope de HIV/metabolismo , Repetição Terminal Longa de HIV/genética , HIV-1 , Transcrição Gênica , Células Cultivadas , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Citoplasma/metabolismo , Retículo Endoplasmático/metabolismo , Regulação Viral da Expressão Gênica , Produtos do Gene gag/metabolismo , Produtos do Gene tat/genética , Genes gag , Proteína gp41 do Envelope de HIV/genética , Humanos , Imunoprecipitação , Provírus , Saccharomyces cerevisiae , Ativação Transcricional , Técnicas do Sistema de Duplo-Híbrido , Replicação Viral , Produtos do Gene tat do Vírus da Imunodeficiência HumanaRESUMO
The human immunodeficiency virus type 1 (HIV-1) encodes a potent transactivator, Tat, which functions through binding to a short leader RNA, called transactivation responsive element (TAR). Recent studies suggest that Tat activates the HIV-1 long terminal repeat (LTR), mainly by adapting co-activator complexes, such as p300, PCAF and the positive transcription elongation factor P-TEFb, to the promoter. Here, we show that the proto-oncoprotein Hdm2 interacts with Tat and mediates its ubiquitination in vitro and in vivo. In addition, Hdm2 is a positive regulator of Tat-mediated transactivation, indicating that the transcriptional properties of Tat are stimulated by ubiquitination. Fusion of ubiquitin to Tat bypasses the requirement of Hdm2 for efficient transactivation, supporting the notion that ubiquitin has a non-proteolytic function in Tat-mediated transactivation.