RESUMO
The E3 small ubiquitin-like modifier (SUMO) protein ligase protein inhibitor of activated STAT 4 (PIAS4) is a pivotal protein in regulating the TGFß pathway. In this study, we discovered a new protein isoform encoded by KIAA0317, termed fibrosis-inducing E3 ligase 1 (FIEL1), which potently stimulates the TGFß signaling pathway through the site-specific ubiquitination of PIAS4. FIEL1 targets PIAS4 using a double locking mechanism that is facilitated by the kinases PKCζ and GSK3ß. Specifically, PKCζ phosphorylation of PIAS4 and GSK3ß phosphorylation of FIEL1 are both essential for the degradation of PIAS4. FIEL1 protein is highly expressed in lung tissues from patients with idiopathic pulmonary fibrosis (IPF), whereas PIAS4 protein levels are significantly reduced. FIEL1 overexpression significantly increases fibrosis in a bleomycin murine model, whereas FIEL1 knockdown attenuates fibrotic conditions. Further, we developed a first-in-class small molecule inhibitor toward FIEL1 that is highly effective in ameliorating fibrosis in mice. This study provides a basis for IPF therapeutic intervention by modulating PIAS4 protein abundance.
Assuntos
Pulmão/metabolismo , Proteínas Inibidoras de STAT Ativados/metabolismo , Fibrose Pulmonar/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação , Animais , Bleomicina/efeitos adversos , Bleomicina/farmacologia , Linhagem Celular , Modelos Animais de Doenças , Glicogênio Sintase Quinase 3 beta/genética , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Pulmão/patologia , Camundongos , Proteínas de Ligação a Poli-ADP-Ribose , Proteínas Inibidoras de STAT Ativados/genética , Proteína Quinase C/genética , Proteína Quinase C/metabolismo , Proteína Quinase C-theta , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/genética , Fibrose Pulmonar/patologia , Ubiquitina-Proteína Ligases/genéticaRESUMO
Invading pathogens may trigger overactivation of the innate immune system, which results in the release of large amounts of proinflammatory cytokines (cytokine storm) and leads to the development of pulmonary edema, multiorgan failure, and shock. PIAS1 is a multifunctional and potent anti-inflammatory protein that negatively regulates several key inflammatory pathways such as Janus kinase (JAK)-signal transducer and activator of transcription (STAT) and nuclear factor κB (NF-κB). We discovered a ubiquitin E3 ligase, HECTD2, which ubiquitinated and mediated the degradation of PIAS1, thus increasing inflammation in an experimental pneumonia model. We found that GSK3ß phosphorylation of PIAS1 provided a phosphodegron for HECTD2 targeting. We also identified a mislocalized HECTD2 polymorphism, HECTD2(A19P), that was present in 8.5% of the population and functioned to reduce inflammation. This polymorphism prevented HECTD2/PIAS1 nuclear interaction, thus preventing PIAS1 degradation. The HECTD2(A19P) polymorphism was also protective toward acute respiratory distress syndrome (ARDS). We then developed a small-molecule inhibitor, BC-1382, that targeted HECTD2 and attenuated lipopolysaccharide (LPS)- and Pseudomonas aeruginosa-induced lung inflammation. These studies describe an unreported innate immune pathway and suggest that mutation or antagonism of the E3 ligase HECTD2 results in reduced severity of lung inflammation by selectively modulating the abundance of the anti-inflammatory protein PIAS1.