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1.
Cell Commun Signal ; 22(1): 48, 2024 01 17.
Artigo em Inglês | MEDLINE | ID: mdl-38233853

RESUMO

BACKGROUND: Interferon Regulatory Factor 3 (IRF3) is a transcription factor that plays a crucial role in the innate immune response by recognizing and responding to foreign antigens. Recently, its roles in sterile conditions are being studied, as in metabolic and fibrotic diseases. However, the search on the upstream regulator for efficient pharmacological targeting is yet to be fully explored. Here, we show that G protein-coupled receptors (GPCRs) can regulate IRF3 phosphorylation through of GPCR-Gα protein interaction. RESULTS: IRF3 and target genes were strongly associated with fibrosis markers in liver fibrosis patients and models. Conditioned media from MIHA hepatocytes overexpressing IRF3 induced fibrogenic activation of LX-2 hepatic stellate cells (HSCs). In an overexpression library screening using active mutant Gα subunits and Phos-tag immunoblotting, Gαs was found out to strongly phosphorylate IRF3. Stimulation of Gαs by glucagon or epinephrine or by Gαs-specific designed GPCR phosphorylated IRF3. Protein kinase A (PKA) signaling was primarily responsible for IRF3 phosphorylation and Interleukin 33 (IL-33) expression downstream of Gαs. PKA phosphorylated IRF3 on a previously unrecognized residue and did not require reported upstream kinases such as TANK-binding kinase 1 (TBK1). Activation of Gαs signaling by glucagon induced IL-33 production in hepatocytes. Conditioned media from the hepatocytes activated HSCs, as indicated by α-SMA and COL1A1 expression, and this was reversed by pre-treatment of the media with IL-33 neutralizing antibody. CONCLUSIONS: Gαs-coupled GPCR signaling increases IRF3 phosphorylation through cAMP-mediated activation of PKA. This leads to an increase of IL-33 expression, which further contributes to HSC activation. Our findings that hepatocyte GPCR signaling regulates IRF3 to control hepatic stellate cell transdifferentiation provides an insight for understanding the complex intercellular communication during liver fibrosis progression and suggests therapeutic opportunities for the disease. Video Abstract.


Assuntos
Células Estreladas do Fígado , Interleucina-33 , Humanos , Interleucina-33/metabolismo , Fator Regulador 3 de Interferon/metabolismo , Transdiferenciação Celular , Meios de Cultivo Condicionados , Glucagon/metabolismo , Hepatócitos/metabolismo , Cirrose Hepática/metabolismo , Fibrose
2.
Biochem Biophys Res Commun ; 681: 186-193, 2023 11 12.
Artigo em Inglês | MEDLINE | ID: mdl-37783116

RESUMO

Primary cilia are essential cellular antennae that transmit external signals into intracellular responses. These sensory organelles perform crucial tasks in triggering intracellular signaling pathways, including those initiated by G protein-coupled receptors (GPCRs). Given the involvement of GPCRs in serum-induced signaling, we investigated the contribution of ciliary proteins in mitogen perception and cell proliferation. We found that depletion of cilia via IFT88 silencing impaired cell growth and repressed YAP activation against serum and its mitogenic constituents, namely lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P). To identify the key player of serum mitogen signaling, a mutant cell line library with 30 ablated individual ciliary proteins was established and screened based on YAP dephosphorylation and target gene induction. While 9 of them had altered signaling, ablation of IFT38 or IFT144 led to a particularly robust repression of YAP activation upon LPA and S1P. The deficiency of IFT38 and IFT144 attenuated cell proliferation, as corroborated in either 2-dimensional cultures or tumor spheroids. In subcutaneous skin melanoma patients, expression of IFT38 and IFT144 was associated with unfavorable outcomes in overall survival. In conclusion, our study demonstrates the involvement of ciliary proteins in mitogen signaling and identifies the regulatory roles of IFT38 and IFT144 in serum-mediated Hippo pathway signaling and cellular growth.


Assuntos
Mitógenos , Transdução de Sinais , Humanos , Linhagem Celular , Proliferação de Células , Lisofosfolipídeos/farmacologia , Receptores Acoplados a Proteínas G/metabolismo
3.
Cell Death Dis ; 15(1): 51, 2024 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-38225223

RESUMO

Yes-associated protein (YAP) and WW domain-containing transcription regulator protein 1 (WWTR1; also known as TAZ) are the main effectors of the Hippo pathway and their dysregulation contributes to diseases in tissues including the liver. Although mitochondria are capable of transmitting signals to change transcriptomic landscape of diseased hepatocytes, such retrograde signaling and the related nuclear machinery are largely unknown. Here, we show that increased YAP activity is associated with mitochondrial stress during liver injury; and this is required for secondary inflammation, promoting hepatocyte death. Mitochondrial stress inducers robustly promoted YAP/TAZ dephosphorylation, nuclear accumulation, and target gene transcription. RNA sequencing revealed that the majority of mitochondrial stress transcripts required YAP/TAZ. Mechanistically, direct oxidation of RhoA by mitochondrial superoxide was responsible for PP2A-mediated YAP/TAZ dephosphorylation providing a novel physiological input for the Hippo pathway. Hepatocyte-specific Yap/Taz ablation suppressed acetaminophen-induced liver injury and blunted transcriptomic changes associated with the pathology. Our observations uncover unappreciated pathway of mitochondrial stress signaling and reveal YAP/TAZ activation as the mechanistic basis for liver injury progression.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Proteínas de Sinalização YAP , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Fígado/metabolismo , Transdução de Sinais , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo
4.
Biomol Ther (Seoul) ; 31(1): 48-58, 2023 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-36579460

RESUMO

Interferon regulatory factor 3 (IRF3) integrates both immunological and non-immunological inputs to control cell survival and death. Small GTPases are versatile functional switches that lie on the very upstream in signal transduction pathways, of which duration of activation is very transient. The large number of homologous proteins and the requirement for site-directed mutagenesis have hindered attempts to investigate the link between small GTPases and IRF3. Here, we constructed a constitutively active mutant expression library for small GTPase expression using Gibson assembly cloning. Small-scale screening identified multiple GTPases capable of promoting IRF3 phosphorylation. Intriguingly, 27 of 152 GTPases, including ARF1, RHEB, RHEBL1, and RAN, were found to increase IRF3 phosphorylation. Unbiased screening enabled us to investigate the sequence-activity relationship between the GTPases and IRF3. We found that the regulation of IRF3 by small GTPases was dependent on TBK1. Our work reveals the significant contribution of GTPases in IRF3 signaling and the potential role of IRF3 in GTPase function, providing a novel therapeutic approach against diseases with GTPase overexpression or active mutations, such as cancer.

5.
Int J Cardiol ; 124(2): e34-6, 2008 Feb 29.
Artigo em Inglês | MEDLINE | ID: mdl-17360060

RESUMO

We report a patient with a significant residual left-to-right shunt who underwent surgical repair for traumatic VSD. This 13-year-old boy was stabbed with a pair of scissors through his heart. After initial life-saving surgery with lacerated left ventricular wall repair and VSD closure, residual VSD was noted. Six months later, we performed a successful transcatheter closure of the residual VSD with an Amplatzer muscular VSD occluder.


Assuntos
Oclusão com Balão/métodos , Comunicação Interventricular/terapia , Septos Cardíacos/lesões , Ferimentos Perfurantes/complicações , Adolescente , Cateterismo Cardíaco/métodos , Procedimentos Cirúrgicos Cardíacos/métodos , Terapia Combinada , Ecocardiografia Transesofagiana , Seguimentos , Comunicação Interventricular/diagnóstico , Humanos , Escala de Gravidade do Ferimento , Masculino , Medição de Risco , Traumatismos Torácicos/complicações , Traumatismos Torácicos/diagnóstico , Traumatismos Torácicos/cirurgia , Resultado do Tratamento , Ferimentos Perfurantes/diagnóstico
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