RESUMO
Rab proteins belong to the Ras superfamily of small monomeric GTPases. These G proteins are the main controllers of vesicular transport in every tissue, among them, the endometrium. They are in charge of to the functional subcellular compartmentalization and cargo transport between organelles and the plasma membrane. In turn, intracellular trafficking contributes to endometrial changes during the menstrual cycle, secretion to the uterine fluid, and trophoblast implantation; however, few reports analyze the role of Rab proteins in the uterus. In general, Rab proteins control the release of cytokines, growth factors, enzymes, hormones, cell adhesion molecules, and mucus. Further, the secretion of multiple compounds into the uterine cavity is required for successful implantation. Therefore, alterations in Rab-controlled intracellular transport likely impair secretory processes to the uterine fluid that may correlate with abnormal endometrial development and failed reproductive outcomes. Overall, they could explain recurrent miscarriages, female infertility, and/or assisted reproductive failure. Interestingly, estrogen (E2) and progesterone (P) regulate gene expression of Rab proteins involved in secretory pathways. This review aims to gather information regarding the role of Rab proteins and intracellular trafficking in the endometrium during the different menstrual phases, and in the generation of a receptive stage for embryo implantation, modulated by E2 and P. This knowledge might be useful for the development of novel reproductive therapies that overcome low implantation rates of assisted reproductive procedures.
Assuntos
Endométrio/metabolismo , Ciclo Menstrual/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Animais , Endométrio/microbiologia , Endométrio/virologia , Estradiol/metabolismo , Feminino , Interações Hospedeiro-Patógeno , Humanos , Progesterona/metabolismo , Transporte Proteico , Doenças Bacterianas Sexualmente Transmissíveis/metabolismo , Doenças Bacterianas Sexualmente Transmissíveis/microbiologia , Doenças Virais Sexualmente Transmissíveis/metabolismo , Doenças Virais Sexualmente Transmissíveis/virologiaRESUMO
Movement and phagocytosis are clue events in colonisation and invasion of tissues by Entamoeba histolytica, the protozoan causative of human amoebiasis. During phagocytosis, EhRab proteins interact with other functional molecules, conducting them to the precise cellular site. The gene encoding EhrabB is located in the complementary chain of the DNA fragment containing Ehcp112 and Ehadh genes, which encode for the proteins of the EhCPADH complex, involved in phagocytosis. This particular genetic organisation suggests that the three corresponding proteins may be functionally related. Here, we studied the relationship of EhRabB with EhCPADH and actin during phagocytosis. First, we obtained the EhRabB 3D structure to carry out docking analysis to predict the interaction sites involved in the EhRabB protein and the EhCPADH complex contact. By confocal microscopy, transmission electron microscopy, and immunoprecipitation assays, we revealed the interaction among these proteins when they move through different vesicles formed during phagocytosis. The role of the actin cytoskeleton in this event was also confirmed using Latrunculin A to interfere with actin polymerisation. This affected the movement of EhRabB and EhCPADH, as well as the rate of phagocytosis. Mutant trophozoites, silenced in EhrabB gene, evidenced the interaction of this molecule with EhCPADH and strengthened the role of actin during erythrophagocytosis.
Assuntos
Citoesqueleto de Actina/ultraestrutura , Entamoeba histolytica/metabolismo , Fagocitose/genética , Trofozoítos/ultraestrutura , Proteínas rab de Ligação ao GTP/química , Proteínas rab de Ligação ao GTP/metabolismo , Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Actinas/ultraestrutura , Membrana Celular/metabolismo , Membrana Celular/ultraestrutura , Entamoeba histolytica/genética , Entamoeba histolytica/patogenicidade , Entamoeba histolytica/ultraestrutura , Eritrócitos/parasitologia , Eritrócitos/ultraestrutura , Humanos , Microscopia Eletrônica de Transmissão , Simulação de Dinâmica Molecular , Mutação , Proteínas de Protozoários/química , Proteínas de Protozoários/metabolismo , Trofozoítos/efeitos dos fármacos , Trofozoítos/metabolismo , Proteínas rab de Ligação ao GTP/genéticaRESUMO
Chlamydia trachomatis, an obligate intracellular bacterium, intercepts different trafficking pathways of the host cell to acquire essential lipids for its survival and replication, particularly from the Golgi apparatus via a Rab14-mediated transport. Molecular mechanisms underlying how these bacteria manipulate intracellular transport are a matter of intense study. Here, we show that C. trachomatis utilizes Akt/AS160 signaling pathway to promote sphingolipids delivery to the chlamydial inclusion through Rab14-controlled vesicular transport. C. trachomatis provokes Akt phosphorylation along its entire developmental life cycle and recruits phosphorylated Akt (pAkt) to the inclusion membrane. As a consequence, Akt Substrate of 160 kDa (AS160), also known as TBC1D4, a GTPase Activating Protein (GAP) for Rab14, is phosphorylated and therefore inactivated. Phosphorylated AS160 (pAS160) loses its ability to promote GTP hydrolysis, favoring Rab14 binding to GTP. Akt inhibition by an allosteric isoform-specific Akt inhibitor (iAkt) prevents AS160 phosphorylation and reduces Rab14 recruitment to chlamydial inclusions. iAkt further impairs sphingolipids acquisition by C. trachomatis-inclusion and provokes lipid retention at the Golgi apparatus. Consequently, treatment with iAkt decreases chlamydial inclusion size, bacterial multiplication, and infectivity in a dose-dependent manner. Similar results were found in AS160-depleted cells. By electron microscopy, we observed that iAkt generates abnormal bacterial forms as those reported after sphingolipids deprivation or Rab14 silencing. Taken together, our findings indicate that targeting the Akt/AS160/Rab14 axis could constitute a novel strategy to limit chlamydial infections, mainly for those caused by antibiotic-resistant bacteria.
RESUMO
Secreção vesicular de macromoléculas foi demonstrada recentemente em Cryptococcus neoformans levantando a hipótese da possibilidade desse mecanismo de transporte vesicular ocorrer também em ascomicetos. Neste trabalho, analisamos se o fungo Histoplasma capsulatum e outros ascomicetos com importância clínica produzem vesículas e utilizam essas estruturas na secreção de macromoléculas. Microscopia eletrônica de transmissão (MET) evidenciou secreção trans-celular de vesículas na fase leveduriforme. Análises proteômicas e de lipídeos revelaram uma gama de macromoléculas envolvidas em diversos processos celulares, como por exemplo: metabolismo, sinalização e virulência. Os resultados demonstraram que H. capsulatum utiliza-se de um transporte secretório celular vesicular trans-membrana para promover virulência...
MET de sobrenadantes de cultura de Candida albicans, Candida parapsilosis, Sporothrix schenckii, e Saccharomyces cerevisiae evidenciaram a presença de vesículas, confirmando a hipótese de que os ascomicetos similarmente produzem vesículas. Anticorpos presentes em pool de soros de pacientes com histoplasmose, reagiram com moléculas presentes nas vesículas isoladas, ressaltando a possível associação dos produtos vesiculares no processo de patogênese. Nossos resultados corroboram a proposta de que a secreção vesicular é um mecanismo comum em fungos para o transporte de macromoléculas relacionadas à virulência, sendo este um promissor alvo em novas linhas terapêuticas...