RESUMO
Tim17 and Tim23 are the main subunits of the TIM23 complex, one of the two major essential mitochondrial inner-membrane protein translocon machineries (TIMs). No chemical probes that specifically inhibit TIM23-dependent protein import were known to exist. Here we show that the natural product stendomycin, produced by Streptomyces hygroscopicus, is a potent and specific inhibitor of the TIM23 complex in yeast and mammalian cells. Furthermore, stendomycin-mediated blockage of the TIM23 complex does not alter normal processing of the major regulatory mitophagy kinase PINK1, but TIM23 is required to stabilize PINK1 on the outside of mitochondria to initiate mitophagy upon membrane depolarization.
Assuntos
Proteínas Mitocondriais/metabolismo , Peptídeos/farmacologia , Proteínas de Saccharomyces cerevisiae/antagonistas & inibidores , Peptídeos Catiônicos Antimicrobianos , Relação Dose-Resposta a Droga , Células HeLa , Humanos , Proteínas de Membrana Transportadoras/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Proteínas do Complexo de Importação de Proteína Precursora Mitocondrial , Estrutura Molecular , Peptídeos/química , Transporte Proteico/efeitos dos fármacos , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
Diverse protein import pathways into mitochondria use translocons on the outer membrane (TOM) and inner membrane (TIM). We adapted a genetic screen, based on Ura3 mistargeting from mitochondria to the cytosol, to identify small molecules that attenuated protein import. Small molecule mitochondrial import blockers of the Carla Koehler laboratory (MB)-10 inhibited import of substrates that require the TIM23 translocon. Mutational analysis coupled with molecular docking and molecular dynamics modeling revealed that MB-10 binds to a specific pocket in the C-terminal domain of Tim44 of the protein-associated motor (PAM) complex. This region was proposed to anchor Tim44 to the membrane, but biochemical studies with MB-10 show that this region is required for binding to the translocating precursor and binding to mtHsp70 in low ATP conditions. This study also supports a direct role for the PAM complex in the import of substrates that are laterally sorted to the inner membrane, as well as the mitochondrial matrix. Thus, MB-10 is the first small molecule modulator to attenuate PAM complex activity, likely through binding to the C-terminal region of Tim44.
Assuntos
Proteínas de Transporte da Membrana Mitocondrial/antagonistas & inibidores , Proteínas de Saccharomyces cerevisiae/antagonistas & inibidores , Saccharomyces cerevisiae/efeitos dos fármacos , Animais , Sítios de Ligação , Testes Genéticos , Células HeLa , Humanos , Proteínas de Transporte da Membrana Mitocondrial/química , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Proteínas do Complexo de Importação de Proteína Precursora Mitocondrial , Modelos Moleculares , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Neurospora crassa , Transporte Proteico/efeitos dos fármacos , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/metabolismo , Peixe-ZebraRESUMO
OBJECTIVE: Do the 3.5 million US veterans, who primarily utilize private healthcare, have similar burn pit exposure and disease compared to the VA Burn Pit registry? METHODS: This is an online volunteer survey of Gulf War and Post-9/11 veterans. RESULTS: Burn pit exposure had significantly higher odds of extremity numbness, aching pain and burning, asthma, chronic obstructive pulmonary disease, interstitial lung disease, constrictive bronchiolitis, pleuritis, and pulmonary fibrosis. Chi-square did not reveal a difference in burn pit exposure and cancer diagnoses. CONCLUSIONS: These data demonstrate increased risk of neurological symptoms associated with burn pit exposure, which are not covered in the 2022 federal Promise to Address Comprehensive Toxics Act. Additional data will allow for the continued review and consideration for future medical benefits.