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Elucidating the Roles of MCM7 and SHCBP1 in Liver Fibrosis through LC-MS Analysis

ABSTRACT: Liver fibrosis, characterized by the excessive deposition of extracellular matrix, presents a significant challenge due to its intricately complex and poorly understood mechanisms. In this study, we aimed to elucidate the roles of Minichromosome Maintenance Complex Component 7 (MCM7) and SHC SH2-Domain Binding Protein 1 (SHCBP1) in liver fibrosis, focusing on their regulatory networks and interactions. Stable hepatocyte cell lines with overexpression of MCM7 and SHCBP1 were constructed. Through targeted LC-MS analysis of MCM7, significant changes were observed in the protein interaction profiles, . MCM7 was found to interact with SHCBP1, which was further analyzed using LC-MS to detail its role in binding to RACGAP1. This interaction activates the SHCBP1-RACGAP1-STAT3 signaling axis, leading to increased IL11 expression. Elevated IL11 levels contribute to the activation of hepatic stellate cells (HSCs) and the progression of liver fibrosis. Our findings demonstrate that through mass spectrometry, MCM7 is identified as a crucial regulatory factor in liver fibrosis, promoting IL11 expression via the SHCBP1-RACGAP1-STAT3 signaling pathway. Both MCM7 and SHCBP1 involvement in driving HSC activation and fibrosis progression. Overall, our study highlights a mechanistic pathway where MCM7 and SHCBP1 collaborate to regulate IL11 expression, thereby influencing the progression of liver fibrosis. These insights emphasize the critical regulatory roles of MCM7 and SHCBP1, offering potential therapeutic targets for combating fibrotic liver diseases.

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ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Hepatocyte, Cell Culture

SUBMITTER: rui zhou

LAB HEAD: ruizhu lab

PROVIDER: PXD055175 | Pride | 2025-08-09

REPOSITORIES: Pride

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Project description:BACKGROUND & AIMS: Hepatocellular carcinoma (HCC) has a high mortality rate due to the lack of effective treatments and drugs. Arsenic trioxide (As2O3, ATO, arsenious acid), which has been proved to successfully treat acute promyelocytic leukemia (APL), was recently reported to show therapeutic potential in solid tumors including liver cancer. Although the mechanistic effects of ATO in APL are established, its anticancer mechanism of action in HCC is still unclear. METHODS: We established an HCC subcutaneous xenograft and intrahepatic metastasis mouse model. In CSC models, tumorspheres and the flow cytometry analysis of CSC markers together with limiting dilution and serial transplantation were used. We compared mRNA expression profiles of the ATO-treated and control cells with mRNA microarray. The expression of target molecule or a clinical correlation was analyzed by immunohistochemistry staining in tissues from ATO-treated mouse and 76 HCC patients. We generated five stable minichromosome maintenance protein 7 (MCM7)-knockdown and two stable MCM7-overexpression HCC cell lines. The chromatin immunoprecipitation (ChIP) assays, immunoprecipitation (IP) assays, the dual luciferase reporter assays, a green biarsenical labeling reagent (FlAsH-EDT2) and a “competing endogenous RNA” (ceRNA) analysis were used. RESULTS: ATO could inhibit the liver tumor-initiating capacity and distant metastasis and prolongs survival in mice. We then screened and found that ATO downregulates the overexpression of MCM7 which is correlated with the progression and prognosis in HCC patients. Knockdown of MCM7 expression recapitulates the inhibition function of ATO on self-renewal of cancer stem cells (CSCs), while overexpression of MCM7 abolishes the inhibition function of ATO on tumorsphere formation. Most importantly, we revealed that ATO directly binds to the MCM7 protein and disturbs the interaction between serum response factor (SRF) and MCM7, resulting in downregulation of MCM7 transcription. A ceRNA analysis also indicated the alterations of endogenous MCM7-associated gene networks involved in stemness-related signaling pathways and cell differentiation. CONCLUSIONS: Here, for the first time, we report that ATO inhibits liver CSCs through blocking the interaction between SRF/MCM7 and suppressing MCM7 autoregulation activity.

Project description:Recurrent hepatitis C virus (rHCV) is universal post-liver transplantation (LT), with accelerated fibrosis rates compared to non-transplanted patients. rHCV is associated with increased mortality and morbidity post-transplant and is a leading indication for re-transplantation. We hypothesized that miRNA expression profiles from liver grafts can distinguish severity of HCV recurrence and differentiate this from acute cellular rejection (ACR). Methods Using microarrays, we characterized global microRNA (miRNA) expression from patients with slow HCV fibrosis progression (F<2 Ishak), fast HCV fibrosis progression (FM-bM-^IM-%2 Ishak), ACR and non-HCV transplanted patients. Selected miRNA were analysed by quantitative PCR (qPCR) using both liver tissue and serum samples. Results We demonstrated changes in miRNA expression in patients with slow HCV fibrosis progression that were anti-fibrogenic, anti-angiogenic and anti-inflammatory in comparison to patients with fast HCV fibrosis progression. miRNA-146a, miRNA-19a, miRNA- 20a and miRNA-let-7e expression were increased in the slow HCV fibrosis progression group. In addition, comparison of patients with fast HCV progression against patients with ACR identified pro-fibrogenic pathways. qPCR analysis on liver tissue and serum confirmed the up-regulation of miRNAs in the slow HCV fibrosis progression group. Conclusion We demonstrate specific miRNA expression signatures that distinguish rate of progression of HCV recurrence and ACR post M-bM-^@M-^Sliver transplantation. Pathway analysis indicates that specific miRNA may play a regulatory role in these processes. The miRNAs identified may act as potential biomarkers for HCV recurrence post-LT and help distinguish between ACR and recurrent HCV. We compared 29 patients in total; 11 patients with slow HCV fibrosis progression, 9 patients with fast HCV fibrosis progression, 5 patients with acute cellular rejection and 4 HCV negative patients with normal liver histology that acted as controls. RNA was extracted from archived histology samples of the RG and NRG and miRNA expression was analysed using the affymetrix Genechip miRNA 2.0 assays.

			Action	DRS
	MCM7-IP-IgG.raw	Raw
	MCM7-IP-MCM7.raw	Raw
	MCM7-Search.rar	Other
	SHCBP1-IP-IgG.raw	Raw
	SHCBP1-IP-SHCBP1.raw	Raw

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Elucidating the Roles of MCM7 and SHCBP1 in Liver Fibrosis through LC-MS Analysis

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