RESUMO
Classic hereditary hemochromatosis (HH) is an autosomal recessive iron-overload disorder resulting from loss-of-function mutations of the HFE gene. Patients with HH exhibit excessive hepatic iron accumulation that predisposes these patients to liver disease, including the risk for developing liver cancer. Chronic iron overload also poses a risk for the development of metabolic disorders such as obesity, type 2 diabetes, and insulin resistance. We hypothesized that liraglutide, GLP1 receptor agonist, alters iron metabolism while also reducing body weight and glucose tolerance in a mouse model of HH (global HFE knockout, HFE KO) and diet-induced obesity and glucose intolerance. The total body HFE KO and wild-type control mice were fed high-fat diet for 8 weeks. Mice were subdivided into liraglutide and vehicle-treated groups and received daily subcutaneous administration of the respective treatment once daily for 18 weeks. Liraglutide improved glucose tolerance and hepatic lipid markers and reduced body weight in a mouse model of HH, the HFE KO mouse, similar to wild-type controls. Importantly, our data show that liraglutide alters iron metabolism in HFE KO mice, leading to decreased circulating and stored iron levels in HFE KO mice. These observations highlight the potential that GLP1 receptor agonist could be used to reduce iron overload in addition to reducing body weight and improving glucose regulation in HH patients.
Assuntos
Modelos Animais de Doenças , Proteína da Hemocromatose , Hemocromatose , Homeostase , Ferro , Liraglutida , Camundongos Knockout , Animais , Hemocromatose/genética , Hemocromatose/metabolismo , Hemocromatose/tratamento farmacológico , Liraglutida/farmacologia , Liraglutida/uso terapêutico , Ferro/metabolismo , Homeostase/efeitos dos fármacos , Camundongos , Proteína da Hemocromatose/genética , Proteína da Hemocromatose/metabolismo , Fígado/metabolismo , Fígado/efeitos dos fármacos , Masculino , Dieta Hiperlipídica/efeitos adversos , Intolerância à Glucose/metabolismo , Intolerância à Glucose/tratamento farmacológico , Intolerância à Glucose/genética , Obesidade/metabolismo , Obesidade/tratamento farmacológico , Obesidade/genética , Camundongos Endogâmicos C57BL , Peso Corporal/efeitos dos fármacosRESUMO
Peroxisomes import their luminal proteins from the cytosol. Most substrates contain a C-terminal Ser-Lys-Leu (SKL) sequence that is recognized by the receptor Pex5. Pex5 binds to peroxisomes via a docking complex containing Pex14, and recycles back into the cytosol following its mono-ubiquitination at a conserved Cys residue. The mechanism of peroxisome protein import remains incompletely understood. Here, we developed an in vitro import system based on Xenopus egg extracts. Import is dependent on the SKL motif in the substrate and on the presence of Pex5 and Pex14, and is sustained by ATP hydrolysis. A protein lacking an SKL sequence can be coimported, providing strong evidence for import of a folded protein. The conserved cysteine in Pex5 is not essential for import or to clear import sites for subsequent rounds of translocation. This new in vitro assay will be useful for further dissecting the mechanism of peroxisome protein import.
Assuntos
Extratos Celulares/análise , Oócitos/metabolismo , Receptor 1 de Sinal de Orientação para Peroxissomos/metabolismo , Peroxissomos/metabolismo , Proteínas de Xenopus/metabolismo , Xenopus laevis/metabolismo , Animais , Citosol/metabolismo , Feminino , Oócitos/citologia , Receptor 1 de Sinal de Orientação para Peroxissomos/genética , Transporte Proteico , Ubiquitinação , Proteínas de Xenopus/genética , Xenopus laevis/genética , Xenopus laevis/crescimento & desenvolvimentoRESUMO
The Pex1 and Pex6 proteins are members of the AAA family of ATPases and are involved in peroxisome biogenesis. Recently, cryo-electron microscopy structures of the Pex1-Pex6 complex in different nucleotide states have been determined. This Structural Snapshot describes the structural features of the complex and their implications for its function, as well as questions that still await answers.
Assuntos
Adenosina Trifosfatases/química , Adenosina Trifosfatases/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Peroxissomos/enzimologia , ATPases Associadas a Diversas Atividades Celulares , Microscopia Crioeletrônica , Humanos , Modelos Moleculares , Complexos Multienzimáticos/química , Complexos Multienzimáticos/metabolismo , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Subunidades Proteicas , Eletricidade EstáticaRESUMO
Members of the AAA family of ATPases assemble into hexameric double rings and perform vital functions, yet their molecular mechanisms remain poorly understood. Here, we report structures of the Pex1/Pex6 complex; mutations in these proteins frequently cause peroxisomal diseases. The structures were determined in the presence of different nucleotides by cryo-electron microscopy. Models were generated using a computational approach that combines Monte Carlo placement of structurally homologous domains into density maps with energy minimization and refinement protocols. Pex1 and Pex6 alternate in an unprecedented hexameric double ring. Each protein has two N-terminal domains, N1 and N2, structurally related to the single N domains in p97 and N-ethylmaleimide sensitive factor (NSF); N1 of Pex1 is mobile, but the others are packed against the double ring. The N-terminal ATPase domains are inactive, forming a symmetric D1 ring, whereas the C-terminal domains are active, likely in different nucleotide states, and form an asymmetric D2 ring. These results suggest how subunit activity is coordinated and indicate striking similarities between Pex1/Pex6 and p97, supporting the hypothesis that the Pex1/Pex6 complex has a role in peroxisomal protein import analogous to p97 in ER-associated protein degradation.
Assuntos
Adenosina Trifosfatases/química , Proteínas de Membrana/química , Proteínas de Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/química , ATPases Associadas a Diversas Atividades Celulares , Difosfato de Adenosina/química , Trifosfato de Adenosina/análogos & derivados , Trifosfato de Adenosina/química , Cromatografia em Gel , Simulação por Computador , Microscopia Crioeletrônica , Retículo Endoplasmático/química , Hidrólise , Método de Monte Carlo , Proteínas Sensíveis a N-Etilmaleimida/química , Peptídeos/química , Peroxissomos/química , Estrutura Terciária de ProteínaRESUMO
Organophosphorus (OP) based pesticides are known powerful inhibitors of cholinesterases, thus the toxicity of this class of compounds causes serious environmental and human health concerns. We report that benzodipyrido[3,2-a:2',3'-c]phenazine (BDPPZ) and 3,6-dimethylbenzodipyrido-[3,2-a:2',3'-c]phenazine (DM-BDPPZ) provide independent fluorescent and electrochemical signal transductions in the presence of the organophosphorus (OP) pesticides; fenthion, malathion and ethion. The presence of the methyl groups at the 3 and 6 positions in DM-BDPPZ was found to significantly influence the sensor performance. The difference in the fluorescence and electrochemical signals produced by the interaction of the sensor compound with each of the OP pesticides provides a means for differentiating between the three pesticides. Detection limits of 10(-8)M, 10(-9) and 10(-12)M were obtained for fenthion, malathion and ethion, respectively. Due to the high sensitivity and ability to minimize false positives these new sensors will be useful for potential integration for future environmental use.