E4 engages uPAR and enolase-1 and activates urokinase to exert antifibrotic effects

Area of Science:

• Biochemistry
• Cell Biology
• Fibrosis Research

Background:

• Fibroproliferative disorders, including systemic sclerosis (SSc), lack effective treatments and cause significant health issues.
• Previous research showed the endostatin C-terminal domain (E4) can prevent and reverse fibrosis in skin and lungs.

Purpose of the Study:

• To elucidate the mechanism of E4's antifibrotic action.
• To identify E4's cell surface binding partners.

Main Methods:

• Assessed E4's effect on the urokinase pathway (uPA/PAI-1 ratio) and matrix metalloproteinases (MMP-1, MMP-3).
• Evaluated E4's in vivo efficacy using a bleomycin-induced fibrosis model.
• Identified E4 binding proteins using biotinylated-E4 and mass spectrometry.
• Investigated the role of identified binding partners (uPAR, ENO) in E4's antifibrotic effects and TGF-β1 signaling.

Main Results:

• E4 activated the urokinase pathway, increasing the uPA/PAI-1 ratio and significantly boosting MMP-1 and MMP-3 expression and activity.
• In vivo, E4 reversed bleomycin-induced PAI-1 increase and enhanced uPA activity.
• Enolase-1 (ENO) and urokinase plasminogen activator receptor (uPAR) were identified as E4 binding proteins.
• E4's antifibrotic effects were dependent on uPAR, while ENO mediated TGF-β1-dependent and independent fibrotic effects.

Conclusions:

• E4's antifibrotic effect is partly mediated by regulating the urokinase pathway and inducing MMP-1/MMP-3 in a uPAR-dependent manner, promoting extracellular matrix degradation.
• ENO exhibits a novel fibrotic role independent of its glycolytic function.