Wilms' tumor 1-associating protein regulates the stress fiber reservoirs in endothelial cells via KLF2-IQGAP3

Stress fibers, renowned for their contractile properties, function as the adhesive glue between cells or between cells and the underlying substrate, playing a pivotal role in cell morphogenesis. Disruptions in stress fibers have been implicated in the onset and progression of various pathological conditions. Wilms tumor 1-associating protein (WTAP), a core constituent of the N6-methyladenosine (m6A) methyltransferase complex, actively participates in the m6A modification of mRNA, thereby modulating gene expression. We investigated the impact of WTAP on stress fibers and the underlying mechanisms. Our study showed that Phorbol 12-myristate 13-acetate (PMA) rapidly triggers the upregulation of WTAP in endothelial cells (ECs), an effect that was mitigated upon inhibition of PKC, Ca²⁺, or NF-κB signaling pathways. Knockdown of WTAP facilitated stress fiber formation and hindered PMA-induced podosome assembly. Under shear stress, ECs with WTAP knockdown exhibited a heightened tendency to align along the direction of flow, as evidenced by VE-cadherin staining. Through RNA-seq analysis, we identified IQ-motif-containing GTPase-activating protein 3 (IQGAP3) as a key target of WTAP, critical for stress fiber formation in ECs. Luciferase reporter assays unveiled the regulatory role of krüppel-like factor 2 on the IQGAP3 promoter and first intron, modulating its expression levels. KLF2 knockdown partially reversed the WTAP-mediated suppression of stress fiber formation. Additionally, treatment of rat thoracic aortas with PMA led to an increase in WTAP protein levels within the endothelium. Our study underscores the pivotal role of WTAP in modulating the stress fiber pool in ECs, with implications for the pathogenesis of vascular diseases.