RESUMEN
HIV-1 Gag precursor directs virus particle assembly and release. In a search for Gag-interacting proteins that are involved in late stages of the HIV-1 replication cycle, we performed yeast two-hybrid screening against a human cDNA library and identified the non-muscle actin filament cross-linking protein filamin A as a novel Gag binding partner. The 280-kDa filamin A regulates cortical actin network dynamics and participates in the anchoring of membrane proteins to the actin cytoskeleton. Recent studies have shown that filamin A facilitates HIV-1 cell-to-cell transmission by binding to HIV receptors and coreceptors and regulating their clustering on the target cell surface. Here we report a novel role for filamin A in HIV-1 Gag intracellular trafficking. We demonstrate that filamin A interacts with the capsid domain of HIV-1 Gag and that this interaction is involved in particle release in a productive manner. Disruption of this interaction eliminated Gag localization at the plasma membrane and induced Gag accumulation within internal compartments. Moreover, blocking clathrin-dependent endocytic pathways did not relieve the restriction to particle release induced by filamin A depletion. These results suggest that filamin A is involved in the distinct step of the Gag trafficking pathway. The discovery of the Gag-filamin A interaction may provide a new therapeutic target for the treatment of HIV infection.
Asunto(s)
Proteínas Contráctiles/metabolismo , Infecciones por VIH/mortalidad , VIH-1/fisiología , Proteínas de Microfilamentos/metabolismo , Ensamble de Virus/fisiología , Clatrina/genética , Clatrina/metabolismo , Proteínas Contráctiles/genética , Endocitosis/genética , Filaminas , Biblioteca de Genes , Infecciones por VIH/genética , Infecciones por VIH/transmisión , VIH-1/patogenicidad , Células HeLa , Humanos , Proteínas de Microfilamentos/genética , Transporte de Proteínas/genética , Saccharomyces cerevisiae , Técnicas del Sistema de Dos Híbridos , Ensamble de Virus/efectos de los fármacos , Productos del Gen gag del Virus de la Inmunodeficiencia HumanaRESUMEN
OBJECTIVE: To determine the effects of levonorgestrel (LNG) on serum androgens, sex hormone-binding globulin (SHBG), hair shaft diameter, and sexual function in women. DESIGN: Substudy of a prospective randomized double-blind study in women using an LNG SC implant (LNG-SI), who were treated with doxycycline or placebo. SETTING: Medical school department of obstetrics and gynecology. PATIENT(S): Forty women were enrolled; 36 completed the study. INTERVENTION(S): Participants were randomized to doxycycline 20 mg or an identical placebo orally twice a day after LNG-SI insertion. MAIN OUTCOME MEASURE(S): Serum levels of total T (TT), free T (FT), androstenedione (A), dehydroepiandrosterone sulfate (DHEAS), dihydrotestosterone (DHT), androstanediol glucuronide (AG), SHBG, and LNG; hair shaft diameter; and sexual function using the Brief Index of Sexual Function for Women and the Arizona Sexual Experiences Scale were assessed. RESULT(S): Serum TT, A, DHT, DHEAS, and SHBG declined after LNG-SI insertion. No changes were found in FT, AG, hair shaft diameter, or sexual function. Serum LNG correlated with SHBG levels. There were no differences between the placebo and doxycycline groups. CONCLUSION(S): LNG reduced serum TT, A, DHT, DHEAS, and SHBG but had no effect on sexual function or markers of androgen bioactivity.