Jiayu Liu

Endothelial cell (EC) mechanosensing is essential for vascular homeostasis and atherosclerosis development, though the influence of blood flow patterns on EC signaling is not fully understood. Discoidin domain receptor 1 (DDR1), a tyrosine kinase receptor, acts as a mechanosensor linking shear force to endothelial responses. Atheroprotective laminar shear flow induces rapid, transient DDR1 activation and condensation through liquid-liquid phase separation without harmful effects, while atherogenic shear causes delayed, sustained condensation, leading to Yes-associated protein (YAP) activation and downstream pathological signaling. We show that heat shock protein 27 (HSP27) regulates shear-dependent DDR1 condensation and phase transition. Specifically, the interaction between DDR1 and HSP27, mediated by their respective domains, is crucial for DDR1 condensate disassembly. Atherogenic shear, unlike atheroprotective shear, triggers prolonged DDR1-mediated HSP27 phosphorylation, promoting DDR1 gel transition and activating YAP signaling. The DDR1-HSP27 axis is key in endothelial YAP activation and atherogenesis in vivo. Targeting this pathway may offer therapeutic potential for preventing endothelial dysfunction and atherosclerosis.

Arterial stiffening, a major cardiovascular risk factor, is driven by aberrant mechanotransduction in vascular smooth muscle cells (VSMCs), yet the critical mechanoreceptors and underlying mechanisms remain elusive. Here, we identified Ephrin receptor A2 (EphA2) as a significantly upregulated mechanosensitive receptor in stiffened arteries from a 5/6 nephrectomy mouse model. Genetic deletion of Epha2 in VSMCs markedly attenuated arterial stiffening. Utilizing polyacrylamide gels of varying stiffness and in situ stiffening bioclick hydrogels, we demonstrated that matrix stiffening directly induces EphA2 phase separation, forming a biomolecular condensate that serves as a signaling hub to recruit and activate ERK1/2. This leads to phosphorylation of the transcription factor CREB and subsequent upregulation of the pro-remodeling nuclear receptor NR4A3. To translate this discovery, we designed a retro-reversed peptide targeting the intrinsically disordered regions (IDRs) of EphA2, which effectively disrupted phase separation and mitigated VSMCs dysfunction in vitro. Crucially, in vivo delivery of this peptide via VAPG-modified nanoparticles significantly alleviated arterial calcification and stiffening in mice. Our study establishes EphA2 phase separation as a pivotal mechanism in vascular mechanotransduction and unveils a novel EphA2-ERK1/2-NR4A3 signaling axis, thereby presenting a promising therapeutic strategy for combating arterial stiffening by targeting pathological biomolecular condensates.

Cell behavior is intricately influenced by the physical forces and mechanical properties of the cells and their microenvironment. Mechanotransduction plays crucial roles in the maintenance of tissue homeostasis and the development of diseases, yet the precise identity of key mechanosensory molecules and structures has been elusive. Recently, the discoidin domain receptor tyrosine kinase 1 (DDR1) has emerged as a novel mechanosensor that responds to mechanical cues such as extracellular matrix stiffness, shear stress, and cytoskeletal tension. In this review, we provide updated insights into DDR1 mechanosensing and propose future research directions. The identification of mechanosensors such as DDR1 holds promise for unlocking novel approaches for drug development to enable precision therapies that target a spectrum of diseases by modulating the mechanotransduction pathways.

Macrophage antigen presentation is crucial for adaptive immunity and maintaining immune balance, including anti-infection, anti-tumor, and inflammation regulation. However, its role in tumor immunomodulation is less understood compared to macrophage polarization. This study explored how Rnaseh2c ⁺ macrophages influence hepatocellular carcinoma (HCC) progression using in vitro cell models and mouse tumor models. Single-cell RNA sequencing, immunoblotting, immunofluorescence, immunoprecipitation, and flow cytometry analysis were employed to examine RNASEH2C’s impact on macrophage antigen presentation. Our results indicated that Rnaseh2c ⁺ macrophages, which were non-polarized, promoted HCC growth by inhibiting antigen presentation. RNASEH2C facilitated lysosomal degradation of RAI14 by enhancing TRAF3IP1 expression and suppressing the mTOR pathway, with HSC70 and CMTM6 playing opposing roles in RAI14 degradation. RAI14, a skeleton protein, facilitated the macropinocytosis of MHC II molecules and tumor-associated antigen, thus activating Th1 cells in HCC. In conclusion, our study revealed how RNASEH2C mediated RAI14’s lysosomal degradation, offering potential targets and strategies for HCC immunotherapy.

BACKGROUND: Atheroprotective shear stress preserves endothelial barrier function, while atheroprone shear stress enhances endothelial permeability. Yet, the underlying mechanisms through which distinct flow patterns regulate EC integrity remain to be clarified. This study aimed to investigate the involvement of Kindlin-2, a key component of focal adhesion and endothelial adherens junctions crucial for regulating endothelial cell (EC) integrity and vascular stability. METHODS: Mouse models of atherosclerosis in EC-specific Kindlin-2 knockout mice ( Kindlin-2 ^iΔEC ) were used to study the role of Kindlin-2 in atherogenesis. Pulsatile shear (12±4 dynes/cm ² ) or oscillatory shear (0.5±4 dynes/cm ² ) were applied to culture ECs. Live-cell imaging, fluorescence recovery after photobleaching assay, and OptoDroplet assay were used to study the liquid-liquid phase separation (LLPS) of Kindlin-2. Co-immunoprecipitation, mutagenesis, proximity ligation assay, and transendothelial electrical resistance assay were used to explore the underlying mechanism of flow-regulated Kindlin-2 function. RESULTS: We found that Kindlin-2 localization is altered under different flow patterns. Kindlin-2 ^iΔEC mice showed heightened vascular permeability. Kindlin-2 ^iΔEC were bred onto ApoE ^−/− mice to generate Kindlin-2 ^iΔEC ; ApoE ^{− /−} mice, which displayed a significant increase in atherosclerosis lesions. In vitro data showed that in ECs, Kindlin-2 underwent LLPS, a critical process for proper focal adhesion assembly, maturation, and junction formation. Mass spectrometry analysis revealed that oscillatory shear increased arginine methylation of Kindlin-2, catalyzed by PRMT5 (protein arginine methyltransferase 5). Functionally, arginine hypermethylation inhibits Kindlin-2 LLPS, impairing focal adhesion assembly and junction maturation. Notably, we identified R290 of Kindlin-2 as a crucial residue for LLPS and a key site for arginine methylation. Finally, pharmacologically inhibiting arginine methylation reduces EC activation and plaque formation. CONCLUSIONS: Collectively, our study elucidates that mechanical force induces arginine methylation of Kindlin-2, thereby regulating vascular stability through its impact on Kindlin-2 LLPS. Targeting Kindlin-2 arginine methylation emerges as a promising hemodynamic-based strategy for treating vascular disorders and atherosclerosis.