Navegando por Palavras-chave "Actin"

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    Expression of genes encoding smooth muscle contractile proteins in vaginal tissue of women with and without pelvic organ prolapse
    (Wiley-Blackwell, 2012-01-01) Bortolini, Maria Augusta Tezelli [UNIFESP]; Shynlova, Oksana; Drutz, Harold P.; Castro, Rodrigo de Aquino [UNIFESP]; Girão, Manoel João Batista Castello [UNIFESP]; Lye, Stephen; Alarab, May; Univ Toronto; Universidade Federal de São Paulo (UNIFESP); Mt Sinai Hosp
    Aims We hypothesize that the expression of genes encoding vaginal smooth muscle (SM) contractile proteins is altered in patients with pelvic organ prolapse (POP) and is influenced by age and menopausal status. We aim to analyze the expression of SM-myosin heavy chain (MHY11), caldesmon (CALD1), SM gamma-actin (ACTG2), and tropomyosin (TPM1), in premenopausal and postmenopausal women with advanced POP and asymptomatic controls. Methods: During total hysterectomy we collected anterior vaginal wall biopsy samples from 55 women, 37 premenopausal (23 patients and 14 controls), and 18 postmenopausal women (13 patients and 5 controls). Total mRNA from the tissues was quantified by real-time RT-PCR. Results: MHY11 gene expression was down-regulated in premenopausal POP patients compared to premenopausal controls (fivefold, P = 0.002). in the postmenopausal groups, we observed a sixfold increase in the CALD1 gene expression in POP patients compared to asymptomatic controls (P = 0.03). the gene expression of CALD1, ACTG2, and TPM1 was significantly down-regulated in vaginal tissue of healthy women after menopause (P < 0.05). Conclusion: Dysregulation of the vaginal SM content in POP patients involves alteration of different cellular pathways according to age and menopausal status. Neurourol. Urodynam. 31:109-114, 2012. (C) 2011 Wiley Periodicals, Inc.
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    Papel das GTPases Cdc42, RhoA e Rac1 no citoesqueleto de actina durante a invasão celular pelas formas tripomastigotas metacíclicas e de cultura de tecido de Trypanosoma cruzi
    (Universidade Federal de São Paulo (UNIFESP), 2019-09-26) Bonifacio, Bruno Souza [UNIFESP]; Mortara, Renato Arruda [UNIFESP]; Ferreira, Éden Ramalho de Araujo [UNIFESP]; Bonfim de Melo, Alexis de Sá Ribeiro do [UNIFESP]; http://lattes.cnpq.br/0533784588961610; http://lattes.cnpq.br/4256258817790019; http://lattes.cnpq.br/3754467086294573; http://lattes.cnpq.br/6655678336873160; Universidade Federal de São Paulo (UNIFESP)
    Cell invasion by Trypanosoma cruzi metacyclic trypomastigotes (TMs) and tissue culture trypomastigotes (TCTs) is a complex process involving parasite-host interactions, and the role of the host cell actin cytoskeleton is still controversial in the literature. The GTPases Cdc42, RhoA and Rac1 and their effectors are key regulators of the actin cytoskeleton promoting its polymerization according to cellular stimuli. In this work we aimed to evaluate actin dynamics and the participation of Cdc42, RhoA and Rac1 during cell invasion by T. cruzi trypomastigote forms. Invasion assays with GTPase-depleted (kd) HeLa cells showed reduced TM invasion in the Cdc42-kd and RhoA-kd groups while TCT presented reduced invasion in Cdc42-kd RhoA-kd and Rac1-kd groups. We also performed invasion assays with HeLa cells overexpressing the studied GTPases, in wild type, constitutively active, dominant negative constructs. Our results showed diverse invasion profiles for either TCTs and TMs, varying between increase or reduction of parasite invasion, depending on the GTPase and construct. Additionally, we investigated the distribution of lysosomes in cells depleted for GTPases. We identified augmented spreading of these organelles even in the absence of parasites. Regarding the GTPases, our results suggest the participation of Cdc42 in both rypomastigote forms whereas Rac1 plays a significant role only for TCTs. On the other hand, the RhoA protein is likely to play a negative role in the invasion of TMs and TCTs. We also performed invasion assays with cells pretreated with Cytocalasin D, Latrunculin B, Phaloidin and Jasplakinolide. Our results showed that all toxins inhibited TMs and TCTs invasion. Finally, actin recruitment assays showed in both TCTs and TMs an inconsistent recruitment profiles, in other words, some parasites do recruit while others do not recruit actin during the invasion process. Taken together, our results suggest that actin cytoskeleton plays an indirect role in the invasion of TCTs and TMs, but other mechanisms that might play a role still need to be elucidated.
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    Perturbations in actin dynamics reconfigure protein complexes that modulate GCN2 activity and promote an eIF2 response
    (Company Of Biologists Ltd, 2016) Silva, Richard Cardoso da [UNIFESP]; Sattlegger, Evelyn; Castilho, Beatriz Amaral de [UNIFESP]
    Genetic and pharmacological interventions in yeast and mammalian cells have suggested a cross-talk between the actin cytoskeleton and protein synthesis. Regulation of the activity of the translation initiation factor 2 (eIF2) is a paramount mechanism for cells to rapidly adjust the rate of protein synthesis and to trigger reprogramming of gene expression in response to internal and external cues. Here, we show that disruption of F-actin in mammalian cells inhibits translation in a GCN2-dependent manner, correlating with increased levels of uncharged tRNA. GCN2 activation increased phosphorylation of its substrate eIF2a and the induction of the integrated stress response master regulator, ATF4. GCN2 activation by latrunculin-B is dependent on GCN1 and inhibited by IMPACT. Our data suggest that GCN2 occurs in two different complexes, GCN2-eEF1A and GCN2-GCN1. Depolymerization of F-actin shifts GCN2 to favor the complex with GCN1, concomitant with GCN1 being released from its binding to IMPACT, which is sequestered by G-actin. These events might further contribute to GCN2 activation. Our findings indicate that GCN2 is an important sensor of the state of the actin cytoskeleton.
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    As vias de Cdc42/N­-WASP e Rac1/WAVE2 na dinâmica de actina durante a invasão celular pelos amastigotas extracelulares de Trypanosoma cruzi
    (Universidade Federal de São Paulo (UNIFESP), 2016-11-30) Melo, Alexis de Sá Ribeiro do Bonfim de [UNIFESP]; Mortara, Renato Arruda [UNIFESP]; http://lattes.cnpq.br/3754467086294573; http://lattes.cnpq.br/4256258817790019; Universidade Federal de São Paulo (UNIFESP)
    Host cell invasion by extracellular amastigotes (EA) of Trypanosoma cruzi is highly dependent on actin cytoskeleton of host cells whose regulatory cellular mechanisms are still poorly understood. Cdc42 and Rac1 GTPases are key mediators of the actin cytoskeleton promoting Arp2/3 complex-dependent actin polymerization via activation of their effector proteins, N-WASP and WAVE2, respectively. The aim of this study was to evaluate the participation Cdc42/N-WASP and Rac1/WAVE2 signaling pathways in actin dynamics during EA internazalization in HeLa cells. Using live-cell imaging in confocal microscope it was observed that Cdc42 and Rac1 are recruited to and colocalize with actin during whole period of EA internalisation. GTPases recruitment was sustained in groups expressing active (CA) or inactive (DN) mutant isoforms when compared to native isoforms (WT). When the invasion ability was compared to control groups, the expression of Rac1 CA and Cdc42 WT increased the number of internalized parasites while Rac1 DN and Cdc42 CA expression reduced it. Additionally, the invasion of EAs is inhibited in cells depleted for Rac1 and also recruitment assays using live cells showed delayed invasion despite no effective reduction in amount of polymerized actin at EA invasion sites. For cells depleted for Cdc42 it was observed inhibition of EA internalization in some experiments, but no inhibition in others; live-cell imaging assays also revealed no delay in actin recruitment in this group. In cells depleted for N-WASP and WAVE2 proteins it was also observed inhibition and delay in the internalization without reduction in actin polymerization. Both proteins were also recruited to and colocalized with actin in EA invasion sites. Finally, depletion of four proteins studied did not affect the density or morphology of membrane projections mobilized by AEs as observed by scanning electron microscopy. The overall result confirms the participation of Cdc42/N-WASP and Rac1/WAVE2 signaling pathways in actin dynamics during EA host cell invasion and encourage the study of other proteins possibly cooperating in these pathways.