Kindlin-2 Phase Separation in Response to Flow Controls Vascular Stability.

Area of Science:

• Biochemistry
• Cell Biology
• Vascular Biology

Background:

• Endothelial barrier function is regulated by shear stress, with atheroprotective and atheroprone patterns influencing endothelial cell (EC) integrity.
• The precise mechanisms by which distinct blood flow patterns modulate EC integrity remain incompletely understood.
• Kindlin-2, a focal adhesion and adherens junction protein, is critical for EC integrity and vascular stability.

Purpose of the Study:

• To investigate the role of Kindlin-2 in regulating endothelial cell integrity under varying shear stress conditions.
• To elucidate the molecular mechanisms by which mechanical forces influence Kindlin-2 function.
• To explore Kindlin-2's involvement in atherogenesis.

Main Methods:

• Utilized EC-specific Kindlin-2 knockout mouse models and atherosclerosis models (ApoE knockout).
• Applied distinct shear stress patterns (pulsatile and oscillatory) to cultured ECs.
• Employed live-cell imaging, biochemical assays (co-immunoprecipitation, mass spectrometry), and biophysical techniques (OptoDroplet, FRAP) to study Kindlin-2 localization, liquid-liquid phase separation (LLPS), and post-translational modifications.

Main Results:

• Kindlin-2 localization and function are modulated by shear stress patterns.
• Kindlin-2 deficiency in ECs leads to increased vascular permeability and exacerbated atherosclerosis in mice.
• Oscillatory shear induces arginine methylation of Kindlin-2 via PRMT5, inhibiting its LLPS, impairing focal adhesion and junction maturation.
• Pharmacological inhibition of arginine methylation reduces EC activation and atherosclerotic plaque formation.

Conclusions:

• Mechanical forces regulate vascular stability by inducing arginine methylation of Kindlin-2, which affects its LLPS.
• Kindlin-2's arginine methylation is a key mediator of flow-dependent EC integrity.
• Targeting Kindlin-2 arginine methylation presents a potential hemodynamic-based therapeutic strategy for vascular disorders and atherosclerosis.