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1.
Microb Biotechnol ; 2021 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-33533574

RESUMO

Whole-cell bioconversion of technical lignins using Pseudomonas putida strains overexpressing amine transaminases (ATAs) has the potential to become an eco-efficient route to produce phenolic amines. Here, a novel cell growth-based screening method to evaluate the in vivo activity of recombinant ATAs towards vanillylamine in P. putida KT2440 was developed. It allowed the identification of the native enzyme Pp-SpuC-II and ATA from Chromobacterium violaceum (Cv-ATA) as highly active towards vanillylamine in vivo. Overexpression of Pp-SpuC-II and Cv-ATA in the strain GN442ΔPP_2426, previously engineered for reduced vanillin assimilation, resulted in 94- and 92-fold increased specific transaminase activity, respectively. Whole-cell bioconversion of vanillin yielded 0.70 ± 0.20 mM and 0.92 ± 0.30 mM vanillylamine, for Pp-SpuC-II and Cv-ATA, respectively. Still, amine production was limited by a substantial re-assimilation of the product and formation of the by-products vanillic acid and vanillyl alcohol. Concomitant overexpression of Cv-ATA and alanine dehydrogenase from Bacillus subtilis increased the production of vanillylamine with ammonium as the only nitrogen source and a reduction in the amount of amine product re-assimilation. Identification and deletion of additional native genes encoding oxidoreductases acting on vanillin are crucial engineering targets for further improvement.

2.
Sex Reprod Healthc ; 28: 100596, 2021 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-33550052

RESUMO

BACKGROUND: It is well established that migrants underutilise sexual and reproductive health (SRH) services for structural and socio-cultural reasons. Sex education at Swedish schools is compulsory and an important part of Swedish upbringing, yet little is known about how migrants perceive this. This study examined migrant parents' views on sex education that their children receive at Swedish schools. METHODS: This is a qualitative study using 14 focus group discussions (74 = n) with Arabic speaking migrant parents attending Swedish integration courses. Qualitative data analysis was used following Saldana's coding method. RESULTS: Migrants' perceptions about sex education provided at Swedish schools are influenced by their home countries, where most participants received no sex education at schools and very little sex education at home. Therefore, values about sex in home countries and Sweden are often contrasting. Consequently, migrant parents are concerned about sex education that their children receive at Swedish schools. There are varied interpretations of sex education, concern over the content and methods taught, and there are numerous assumptions about potential negative effects of sex education. CONCLUSION: The study echoes findings on intergenerational challenges in migrant families surrounding the topic of gender and sex, often brought on by discussions about sex education at schools. Innovative approaches are needed to support migrant parents as part of Sweden's effort to strengthen effectiveness and inclusiveness of sex education.

4.
Syst Rev ; 10(1): 37, 2021 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-33485393

RESUMO

BACKGROUND: Global responses to the COVID-19 pandemic have exposed and exacerbated existing socioeconomic and health inequities that disproportionately affect the sexual health and well-being of many populations, including people of color, ethnic minority groups, women, and sexual and gender minority populations. Although there have been several reviews published on COVID-19 and health disparities across various populations, none has focused on sexual health. We plan to conduct a scoping review that seeks to fill several of the gaps in the current knowledge of sexual health in the COVID-19 era. METHODS: A scoping review focusing on sexual health and COVID-19 will be conducted. We will search (from January 2020 onwards) CINAHL, Africa-Wide Information, Web of Science Core Collection, Embase, Gender Studies Database, Gender Watch, Global Health, WHO Global Literature on Coronavirus Disease Database, WHO Global Index Medicus, PsycINFO, MEDLINE, and Sociological Abstracts. Grey literature will be identified using Disaster Lit, Google Scholar, governmental websites, and clinical trials registries (e.g., ClinicalTrial.gov , World Health Organization, International Clinical Trials Registry Platform, and International Standard Randomized Controlled Trial Number Registry). Study selection will conform to the Joanna Briggs Institute Reviewers' Manual 2015 Methodology for JBI Scoping Reviews. Only English language, original studies will be considered for inclusion. Two reviewers will independently screen all citations, full-text articles, and abstract data. A narrative summary of findings will be conducted. Data analysis will involve quantitative (e.g., frequencies) and qualitative (e.g., content and thematic analysis) methods. DISCUSSION: Original research is urgently needed to mitigate the risks of COVID-19 on sexual health. The planned scoping review will help to address this gap. SYSTEMATIC REVIEW REGISTRATIONS: Systematic Review Registration: Open Science Framework osf/io/PRX8E.


Assuntos
/enzimologia , Grupos Étnicos , Pandemias , Saúde Sexual , Minorias Sexuais e de Gênero , /psicologia , Feminino , Saúde Global , Humanos , Masculino , Grupos Minoritários
5.
Methods Mol Biol ; 2169: 119-127, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32548824

RESUMO

The dynamic assembly of proteins at the membrane interphase is key to many cell biological processes such as the generation and stabilization of caveolae at the cell surface via coat proteins. The liposome co-sedimentation assay has been widely used for studies of protein and lipid interactions and has provided important information about binding mechanisms, lipid-binding specificity, and curvature preference of proteins. Here, we describe this technique in detail and how it can be used as a tool to address the membrane-binding ability and lipid specificity of caveolae-associated proteins.

6.
Elife ; 92020 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-32364496

RESUMO

Caveolae are bulb-shaped invaginations of the plasma membrane (PM) that undergo scission and fusion at the cell surface and are enriched in specific lipids. However, the influence of lipid composition on caveolae surface stability is not well described or understood. Accordingly, we inserted specific lipids into the cell PM via membrane fusion and studied their acute effects on caveolae dynamics. We demonstrate that sphingomyelin stabilizes caveolae to the cell surface, whereas cholesterol and glycosphingolipids drive caveolae scission from the PM. Although all three lipids accumulated specifically in caveolae, cholesterol and sphingomyelin were actively sequestered, whereas glycosphingolipids diffused freely. The ATPase EHD2 restricts lipid diffusion and counteracts lipid-induced scission. We propose that specific lipid accumulation in caveolae generates an intrinsically unstable domain prone to scission if not restrained by EHD2 at the caveolae neck. This work provides a mechanistic link between caveolae and their ability to sense the PM lipid composition.

7.
Reprod Health ; 17(1): 67, 2020 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-32434523

RESUMO

BACKGROUND: Adolescent pregnancies are persistently high among refugees. The pregnancies have been attributed to low contraceptive use in this population. The aim of this study was to determine the prevalence and factors associated with modern contraceptive use among female refugee adolescents in northern Uganda. METHODS: This was a cross sectional study using both descriptive and analytical techniques. The study was carried out in Palabek refugee settlement in Northern Uganda from May to July 2019. A total of 839 refugee adolescents who were sexually active or in-union were consecutively enrolled. Interviewer administered questionnaires were used for data collection. RESULTS: Modern contraceptive prevalence was 8.7% (95% CI: 7.0 to 10.8). The injectable was the most commonly used modern contraceptive method [42.5% (95% CI: 31.5 to 54.3)], and most of the participants had used the contraceptives for 6 months or less (59.7%). Reasons for not using modern contraceptives included fear of side effects (39.3%), partner prohibition (16.4%), and the desire to become pregnant (7.0%). Participants who were married (OR = 0.11, 95% CI: 0.04 to 0.35, p < 0.001), cohabiting (OR = 0.43, 95% CI: 0.20 to 0.93, p = 0.032) or having an older partner (OR = 0.93, 95% CI: 0.86 to 0.99, p = 0.046) were less likely to use modern contraceptives. CONCLUSION: Modern contraceptive use among female refugee adolescents was very low, and few reported a desire to become pregnant, leaving them vulnerable to unplanned pregnancies. Least likely to use modern contraceptives were participants who were married/cohabiting and those having older partners implying a gender power imbalance in fertility decision making. There is an urgent need for innovations to address the gender and power imbalances within relationships, which could shape fertility decision-making and increase modern contraceptive use among refugee adolescents.

8.
Proc Natl Acad Sci U S A ; 117(13): 7471-7481, 2020 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-32170013

RESUMO

Eps15-homology domain containing protein 2 (EHD2) is a dynamin-related ATPase located at the neck of caveolae, but its physiological function has remained unclear. Here, we found that global genetic ablation of EHD2 in mice leads to increased lipid droplet size in fat tissue. This organismic phenotype was paralleled at the cellular level by increased fatty acid uptake via a caveolae- and CD36-dependent pathway that also involves dynamin. Concomitantly, elevated numbers of detached caveolae were found in brown and white adipose tissue lacking EHD2, and increased caveolar mobility in mouse embryonic fibroblasts. EHD2 expression itself was down-regulated in the visceral fat of two obese mouse models and obese patients. Our data suggest that EHD2 controls a cell-autonomous, caveolae-dependent fatty acid uptake pathway and imply that low EHD2 expression levels are linked to obesity.


Assuntos
Proteínas de Transporte/metabolismo , Cavéolas/metabolismo , Ácidos Graxos/metabolismo , Animais , Transporte Biológico , Membrana Celular/metabolismo , Células HeLa , Humanos , Gotículas Lipídicas/metabolismo , Metabolismo dos Lipídeos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
9.
Biochem Soc Trans ; 48(1): 155-163, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-32049332

RESUMO

Caveolae are small Ω-shaped invaginations of the plasma membrane that play important roles in mechanosensing, lipid homeostasis and signaling. Their typical morphology is characterized by a membrane funnel connecting a spherical bulb to the membrane. Membrane funnels (commonly known as necks and pores) are frequently observed as transient states during fusion and fission of membrane vesicles in cells. However, caveolae display atypical dynamics where the membrane funnel can be stabilized over an extended period of time, resulting in cell surface constrained caveolae. In addition, caveolae are also known to undergo flattening as well as short-range cycles of fission and fusion with the membrane, requiring that the membrane funnel closes or opens up, respectively. This mini-review considers the transition between these different states and highlights the role of the protein and lipid components that have been identified to control the balance between surface association and release of caveolae.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Transporte/metabolismo , Cavéolas/metabolismo , Caveolina 1/metabolismo , Filaminas/metabolismo , Lipídeos de Membrana/metabolismo , Animais , Proteínas de Transporte/genética , Células HeLa , Humanos , Camundongos , Camundongos Knockout , Transdução de Sinais
10.
J Colloid Interface Sci ; 553: 820-833, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31284226

RESUMO

Despite increasing interests in non-lamellar liquid crystalline dispersions, such as hexosomes, for drug delivery, little is known about their interactions with cells and mechanism of cell entry. Here we examine the cellular uptake of hexosomes based on phytantriol and mannide monooleate by HeLa cells using live cell microscopy in comparison to conventional liposomes. To investigate the importance of specific endocytosis pathways upon particle internalization, we silenced regulatory proteins of major endocytosis pathways using short interfering RNA. While endocytosis plays a significant role in liposome internalization, hexosomes are not taken up via endocytosis but through a mechanism that is dependent on cell membrane tension. Biophysical studies using biomembrane models highlighted that hexosomes have a high affinity for membranes and an ability to disrupt lipid layers. Our data suggest that direct biomechanical interactions of hexosomes with membrane lipids play a crucial role and that the unique morphology of hexosomes is vital for their membrane activity. Based on these results, we propose a mechanism, where hexosomes destabilize the bilayer, allowing them to "phase through" the membrane. Understanding parameters that influence the uptake of hexosomes is critical to establish them as carrier systems that can potentially deliver therapeutics efficiently to intracellular sites of action.


Assuntos
Coloides/metabolismo , Endocitose , Álcoois Graxos/metabolismo , Transporte Biológico , Coloides/síntese química , Coloides/química , Sistemas de Liberação de Medicamentos , Álcoois Graxos/síntese química , Álcoois Graxos/química , Células HeLa , Humanos , Lipossomos/química , Manitol/análogos & derivados , Manitol/síntese química , Manitol/química , Manitol/metabolismo , Ácidos Oleicos/síntese química , Ácidos Oleicos/química , Ácidos Oleicos/metabolismo
11.
J Virol ; 93(18)2019 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-31243132

RESUMO

Flavivirus is a positive-sense, single-stranded RNA viral genus, with members causing severe diseases in humans such as tick-borne encephalitis, yellow fever, and dengue fever. Flaviviruses are known to cause remodeling of intracellular membranes into small cavities, where replication of the viral RNA takes place. Nonstructural (NS) proteins are not part of the virus coat and are thought to participate in the formation of these viral replication compartments (RCs). Here, we used tick-borne encephalitis virus (TBEV) as a model for the flaviviruses and developed a stable human cell line in which the expression of NS proteins can be induced without viral RNA replication. The model system described provides a novel and benign tool for studies of the viral components under controlled expression levels. We show that the expression of six NS proteins is sufficient to induce infection-like dilation of the endoplasmic reticulum (ER) and the formation of RC-like membrane invaginations. The NS proteins form a membrane-associated complex in the ER, and electron tomography reveals that the dilated areas of the ER are closely associated with lipid droplets and mitochondria. We propose that the NS proteins drive the remodeling of ER membranes and that viral RNA, RNA replication, viral polymerase, and TBEV structural proteins are not required.IMPORTANCE TBEV infection causes a broad spectrum of symptoms, ranging from mild fever to severe encephalitis. Similar to other flaviviruses, TBEV exploits intracellular membranes to build RCs for viral replication. The viral NS proteins have been suggested to be involved in this process; however, the mechanism of RC formation and the roles of individual NS proteins remain unclear. To study how TBEV induces membrane remodeling, we developed an inducible stable cell system expressing the TBEV NS polyprotein in the absence of viral RNA replication. Using this system, we were able to reproduce RC-like vesicles that resembled the RCs formed in flavivirus-infected cells, in terms of morphology and size. This cell system is a robust tool to facilitate studies of flavivirus RC formation and is an ideal model for the screening of antiviral agents at a lower biosafety level.


Assuntos
Vírus da Encefalite Transmitidos por Carrapatos/metabolismo , Proteínas não Estruturais Virais/metabolismo , Estruturas Virais/metabolismo , Vírus da Encefalite Transmitidos por Carrapatos/genética , Encefalite Transmitida por Carrapatos/metabolismo , Encefalite Transmitida por Carrapatos/virologia , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/virologia , Flavivirus/genética , Flavivirus/metabolismo , Expressão Gênica/genética , Regulação Viral da Expressão Gênica/genética , Células HeLa , Humanos , Modelos Biológicos , RNA Viral/genética , Proteínas não Estruturais Virais/fisiologia , Estruturas Virais/fisiologia , Replicação Viral/fisiologia
12.
Sex Reprod Healthc ; 19: 64-70, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30928137

RESUMO

OBJECTIVE: Poor sexual and reproductive health (SRH) among immigrant women is often related to limited access, or suboptimal use of healthcare services. This study investigates the knowledge about and use of sexual and reproductive healthcare services among immigrant women in Sweden. METHOD: A cross-sectional study of 288 immigrant women. A structured questionnaire was distributed among immigrants speaking Arabic, Dari, Somali or English registered at Swedish language schools for immigrants. Data collection took place in 19 strategically selected schools in Sweden. Descriptive statistics, chi-square tests, and logistic regressions were used for the analysis. RESULTS: About one-third of the immigrant women reported lack of knowledge of where to go for contraceptive counselling. Experiencing lack of emotional social support and not having had children was associated with this lack of knowledge. An even higher proportion (56%) lacked knowledge of where to go to be HIV tested, and this was associated with not having participated in a health examination. Almost 25% stated that their culture kept them back from using contraception. CONCLUSION: Lack of knowledge of where to turn for contraceptive counselling and HIV testing among immigrant women participating in Swedish language schools for immigrants could be considered as a missed opportunity, as all citizens in Sweden have free access to these services. New health policies and strategies should aim to increase knowledge of SRH services among immigrants. Swedish language schools could play an important role in increasing knowledge of SRH-related information as many new immigrants become students during their first years in Sweden.


Assuntos
Comportamento Contraceptivo/etnologia , Emigrantes e Imigrantes/psicologia , Emigrantes e Imigrantes/estatística & dados numéricos , Infecções por HIV/diagnóstico , Conhecimentos, Atitudes e Prática em Saúde , Serviços de Saúde Reprodutiva/estatística & dados numéricos , Adolescente , Adulto , Afeganistão/etnologia , Aconselhamento , Estudos Transversais , Características da Família , Feminino , Humanos , Iraque/etnologia , Idioma , Pessoa de Meia-Idade , Instituições Acadêmicas , Apoio Social , Somália/etnologia , Inquéritos e Questionários , Suécia , Síria/etnologia , Adulto Jovem
13.
Artigo em Inglês | MEDLINE | ID: mdl-30665889

RESUMO

INTRODUCTION: The objective of this study was to compare ever-in life contraception use, use of contraception at current conception, and planned use of contraception after an induced abortion, among three groups of women: migrants, second-generation migrants and non-migrant women, and to compare the types of contraception methods used and intended for future use among the three groups of women. METHODS: The cross-sectional study administered a questionnaire face-to-face to women aged 18 years and older who were seeking abortion care at one of six abortion clinics in Stockholm County from January to April 2015. RESULTS: The analysis included 637 women. Migrants and second-generation migrants were less likely to have used contraception historically, at the time of the current conception, and to plan to use contraception after their induced abortion compared with non-migrant women. Historically, non-migrants had used pills (89%) and withdrawal (24%) while migrants had used the copper intrauterine device (24%) to a higher extent compared to the other two groups of women. Both the migrants (65%) and second-generation migrants (61%) were more likely than the non-migrants (48%) to be planning to use long-acting reversible contraception. CONCLUSIONS: Lower proportions of contraception use were found in migrants and second-generation migrants than in non-migrants. In addition, there were significant differences in the types of contraception methods used historically and intended for future use.

14.
Health Res Policy Syst ; 16(1): 123, 2018 Dec 17.
Artigo em Inglês | MEDLINE | ID: mdl-30558612

RESUMO

BACKGROUND: The uptake of findings from sexual and reproductive health and rights research into policy-making remains a complex and non-linear process. Different models of research utilisation and guidelines to maximise this in policy-making exist, however, challenges still remain for researchers to improve uptake of their research findings and for policy-makers to use research evidence in their work. METHODS: A participatory workshop with researchers was organised in November 2017 by the Academic Network for Sexual and Reproductive Health and Rights Policy (ANSER) to address this gap. ANSER is a consortium of experienced researchers, some of whom have policy-making experience, working on sexual and reproductive health and rights issues across 16 countries and 5 continents. The experiential learning cycle was used to guide the workshop discussions based on case studies and to encourage participants to focus on key lessons learned. Workshop findings were thematically analysed using specific stages from Hanney et al.'s (Health Res Policy Syst 1:2, 2003) framework on the place of policy-making in the stages of assessment of research utilisation and outcomes. RESULTS: The workshop identified key strategies for translating research into policy, including joint agenda-setting between researchers and policy-makers, as well as building trust and partnerships with different stakeholders. These were linked to stages within Hanney et al.'s framework as opportunities for engaging with policy-makers to ensure uptake of research findings. CONCLUSION: The engagement of stakeholders during the research development and implementation phases, especially at strategic moments, has a positive impact on uptake of research findings. The strategies and stages described in this paper can be applied to improve utilisation of research findings into policy development and implementation globally.


Assuntos
Assistência à Saúde , Política de Saúde , Pesquisa sobre Serviços de Saúde , Formulação de Políticas , Saúde Reprodutiva , Saúde Sexual , Pesquisa Médica Translacional , Pessoal Administrativo , Humanos , Direitos Sexuais e Reprodutivos , Pesquisadores , Participação dos Interessados
15.
Biol Open ; 7(10)2018 Oct 22.
Artigo em Inglês | MEDLINE | ID: mdl-30254077

RESUMO

Endocytic mechanisms have been suggested to be important for plasma membrane repair in response to pore-forming toxins such as listeriolysin O (LLO), which form membrane pores that disrupt cellular homeostasis. Yet, little is known about the specific role of distinct endocytic machineries in this process. Here, we have addressed the importance of key endocytic pathways and developed reporter systems for real-time imaging of the endocytic response to LLO pore formation. We found that loss of clathrin-independent endocytic pathways negatively influenced the efficiency of membrane repair. However, we did not detect any increased activity of these pathways, or co-localisation with the toxin or markers of membrane repair, suggesting that they were not directly involved in removal of LLO pores from the plasma membrane. In fact, markers of clathrin-independent carriers (CLICs) were rapidly disassembled in the acute phase of membrane damage due to Ca2+ influx, followed by a reassembly about 2 min after pore formation. We propose that these endocytic mechanisms might influence membrane repair by regulating the plasma membrane composition and tension, but not via direct internalisation of LLO pores.

16.
J Cell Biol ; 216(11): 3745-3765, 2017 11 06.
Artigo em Inglês | MEDLINE | ID: mdl-28923975

RESUMO

The conditional use of actin during clathrin-mediated endocytosis in mammalian cells suggests that the cell controls whether and how actin is used. Using a combination of biochemical reconstitution and mammalian cell culture, we elucidate a mechanism by which the coincidence of PI(4,5)P2 and PI(3)P in a curved vesicle triggers actin polymerization. At clathrin-coated pits, PI(3)P is produced by the INPP4A hydrolysis of PI(3,4)P2, and this is necessary for actin-driven endocytosis. Both Cdc42⋅guanosine triphosphate and SNX9 activate N-WASP-WIP- and Arp2/3-mediated actin nucleation. Membrane curvature, PI(4,5)P2, and PI(3)P signals are needed for SNX9 assembly via its PX-BAR domain, whereas signaling through Cdc42 is activated by PI(4,5)P2 alone. INPP4A activity is stimulated by high membrane curvature and synergizes with SNX9 BAR domain binding in a process we call curvature cascade amplification. We show that the SNX9-driven actin comets that arise on human disease-associated oculocerebrorenal syndrome of Lowe (OCRL) deficiencies are reduced by inhibiting PI(3)P production, suggesting PI(3)P kinase inhibitors as a therapeutic strategy in Lowe syndrome.


Assuntos
Actinas/metabolismo , Vesículas Revestidas por Clatrina/metabolismo , Clatrina/metabolismo , Invaginações Revestidas da Membrana Celular/metabolismo , Endocitose , Fosfatidilinositóis/metabolismo , Complexo 2-3 de Proteínas Relacionadas à Actina/genética , Complexo 2-3 de Proteínas Relacionadas à Actina/metabolismo , Animais , Sistemas CRISPR-Cas , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/metabolismo , Células HeLa , Humanos , Hidrólise , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Síndrome Oculocerebrorrenal/genética , Síndrome Oculocerebrorrenal/metabolismo , Fosfatidilinositol 4,5-Difosfato/metabolismo , Fosfatos de Fosfatidilinositol/metabolismo , Monoéster Fosfórico Hidrolases/genética , Monoéster Fosfórico Hidrolases/metabolismo , Multimerização Proteica , Interferência de RNA , Epitélio Pigmentado da Retina/metabolismo , Transdução de Sinais , Nexinas de Classificação/genética , Nexinas de Classificação/metabolismo , Fatores de Tempo , Transfecção , Proteína Neuronal da Síndrome de Wiskott-Aldrich/genética , Proteína Neuronal da Síndrome de Wiskott-Aldrich/metabolismo , Xenopus laevis , Proteína cdc42 de Ligação ao GTP/genética , Proteína cdc42 de Ligação ao GTP/metabolismo
17.
Cell Rep ; 20(8): 1893-1905, 2017 Aug 22.
Artigo em Inglês | MEDLINE | ID: mdl-28834752

RESUMO

Cellular blebbing, caused by local alterations in cell-surface tension, has been shown to increase the invasiveness of cancer cells. However, the regulatory mechanisms balancing cell-surface dynamics and bleb formation remain elusive. Here, we show that an acute reduction in cell volume activates clathrin-independent endocytosis. Hence, a decrease in surface tension is buffered by the internalization of the plasma membrane (PM) lipid bilayer. Membrane invagination and endocytosis are driven by the tension-mediated recruitment of the membrane sculpting and GTPase-activating protein GRAF1 (GTPase regulator associated with focal adhesion kinase-1) to the PM. Disruption of this regulation by depleting cells of GRAF1 or mutating key phosphatidylinositol-interacting amino acids in the protein results in increased cellular blebbing and promotes the 3D motility of cancer cells. Our data support a role for clathrin-independent endocytic machinery in balancing membrane tension, which clarifies the previously reported role of GRAF1 as a tumor suppressor.


Assuntos
Clatrina/metabolismo , Endocitose/fisiologia , Pseudópodes/fisiologia , Fenômenos Biológicos , Humanos , Invasividade Neoplásica
18.
Qual Health Res ; 27(10): 1518-1528, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28728532

RESUMO

In this article, we explore how reproductive health care providers in Sweden, a country often described as one of the most gender-equal countries in the world, incorporate gender equality ideals in multicultural contraceptive counseling. In the tension between gender equality promotion on one hand and respect for cultural diversity and individualized care on the other, we will demonstrate that values of gender equality were often given priority. This is not necessarily undesirable. Nevertheless, our proposal is that the gender equality ideology may inhibit providers' ability to think differently about issues at stake in contraceptive counseling, which may negatively influence women's possibilities to obtain adequate support. At the end of the article, we suggest how health care providers' reflexivity might be used as a working tool for increased awareness about the taken-for-granted cultural norms that exist in their clinical milieu.


Assuntos
Atitude do Pessoal de Saúde , Anticoncepção/psicologia , Aconselhamento/métodos , Serviços de Planejamento Familiar/métodos , Pessoal de Saúde/psicologia , Adulto , Feminino , Humanos , Masculino , Fatores Sexuais , Suécia
19.
Proc Natl Acad Sci U S A ; 114(22): 5629-5634, 2017 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-28228524

RESUMO

Eps15 (epidermal growth factor receptor pathway substrate 15)-homology domain containing proteins (EHDs) comprise a family of dynamin-related mechano-chemical ATPases involved in cellular membrane trafficking. Previous studies have revealed the structure of the EHD2 dimer, but the molecular mechanisms of membrane recruitment and assembly have remained obscure. Here, we determined the crystal structure of an amino-terminally truncated EHD4 dimer. Compared with the EHD2 structure, the helical domains are 50° rotated relative to the GTPase domain. Using electron paramagnetic spin resonance (EPR), we show that this rotation aligns the two membrane-binding regions in the helical domain toward the lipid bilayer, allowing membrane interaction. A loop rearrangement in GTPase domain creates a new interface for oligomer formation. Our results suggest that the EHD4 structure represents the active EHD conformation, whereas the EHD2 structure is autoinhibited, and reveal a complex series of domain rearrangements accompanying activation. A comparison with other peripheral membrane proteins elucidates common and specific features of this activation mechanism.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adenosina Trifosfatases/metabolismo , Proteínas de Transporte/metabolismo , Membrana Celular/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas Nucleares/metabolismo , Sequência de Aminoácidos/genética , Linhagem Celular Tumoral , Cristalografia por Raios X , Ativação Enzimática/fisiologia , Células HeLa , Humanos , Ligação Proteica , Domínios Proteicos/fisiologia , Multimerização Proteica , Transporte Proteico/fisiologia
20.
Proc Natl Acad Sci U S A ; 114(22): E4360-E4369, 2017 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-28223496

RESUMO

The EH-domain-containing protein 2 (EHD2) is a dynamin-related ATPase that confines caveolae to the cell surface by restricting the scission and subsequent endocytosis of these membrane pits. For this, EHD2 is thought to first bind to the membrane, then to oligomerize, and finally to detach, in a stringently regulated mechanistic cycle. It is still unclear how ATP is used in this process and whether membrane binding is coupled to conformational changes in the protein. Here, we show that the regulatory N-terminal residues and the EH domain keep the EHD2 dimer in an autoinhibited conformation in solution. By significantly advancing the use of infrared reflection-absorption spectroscopy, we demonstrate that EHD2 adopts an open conformation by tilting the helical domains upon membrane binding. We show that ATP binding enables partial insertion of EHD2 into the membrane, where G-domain-mediated oligomerization occurs. ATP hydrolysis is related to detachment of EHD2 from the membrane. Finally, we demonstrate that the regulation of EHD2 oligomerization in a membrane-bound state is crucial to restrict caveolae dynamics in cells.


Assuntos
Proteínas de Transporte/química , Proteínas de Transporte/metabolismo , Trifosfato de Adenosina/metabolismo , Substituição de Aminoácidos , Animais , Proteínas de Transporte/genética , Cavéolas/metabolismo , Transferência Ressonante de Energia de Fluorescência , Células HeLa , Humanos , Camundongos , Simulação de Dinâmica Molecular , Mutagênese Sítio-Dirigida , Ligação Proteica , Conformação Proteica , Estrutura Quaternária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Espectrofotometria Infravermelho
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