RAF dimers control vascular permeability and cytoskeletal rearrangements at endothelial cell‐cell junctions

Abstract

The endothelium functions as a semipermeable barrier regulating fluid homeostasis, nutrient, and gas supply to the tissue. Endothelial permeability is increased in several pathological conditions including inflammation and tumors; despite its clinical relevance, however, there are no specific therapies preventing vascular leakage. Here, we show that endothelial cell‐restricted ablation of BRAF , a kinase frequently activated in cancer, prevents vascular leaking as well metastatic spread. BRAF regulates endothelial permeability by promoting the cytoskeletal rearrangements necessary for the remodeling of VE ‐Cadherin‐containing endothelial cell–cell junctions and the formation of intercellular gaps. BRAF kinase activity and the ability to form complexes with RAS / RAP 1 and dimers with its paralog RAF 1 are required for proper permeability control, achieved mechanistically by modulating the interaction between RAF 1 and the RHO effector ROK α. Thus, RAF dimerization impinges on RHO pathways to regulate cytoskeletal rearrangements, junctional plasticity, and endothelial permeability. The data advocate the development of RAF dimerization inhibitors, which would combine tumor cell autonomous effect with stabilization of the vasculature and antimetastatic spread.

BRAF, A gatekeeper controlling endothelial permeability

“…This was further supported by the fact that BRAF‐deficient confluent endothelial monolayers did not show radial stress fiber formation—normally formed upon permeability‐inducing stimuli—nor intracellular gaps even after stimulation with VEGF, a potent permeability inducer frequently upregulated in tumors. BRAF‐deficient pMECs were characterized by prominent circumferential actin bundles which correlated with less F‐actin content . These results are in accordance with previous findings in the literature reporting that BRAF KO mouse embryonic fibroblasts have reduced actin stress fibers and F‐actin content .…”

“…A mouse model of BRAF ablation in the endothelial compartment (using a VE-Cadherin-Cre reporter) did not cause any developmental defects and tissue architecture of the major organs tested remained unaffected. Besides, postnatal retinal angiogenesis and adult angiogenesis were normal, as tested via Matrigel plug and sprouting bead assays using BRAF-deficient primary microvessel-derived mouse lung endothelial cells (pMECs) [3]. These results are in agreement with the observation that BRAF is not required for EC survival in embryonic and adult mice [5].…”

“…Consistently with what was found in BRAF KO mouse embryonic fibroblasts , Dorard et al . found reduced phosphorylation of LIMK and its downstream target COFILIN in BRAF‐deficient ECs. However, VEGF signaling upstream of ROKα was unaltered suggesting a roadblock in RHOA signaling at the level of ROKα.…”

“…This mechanism was excluded by Dorard et al . using a MEK inhibitor which dampened ERK activation in wild‐type pMECs but had no impact on BRAF KO pMECs, demonstrating that reduced MEK/ERK activation in BRAF‐deficient pMEC is not the cause of decreased permeability.…”

“…However, such treatments face drug resistance with the onset of side effects due to parallel signaling circuits activation, via yet incompletely understood combinations of RAF paralogs [2]. The article by Dorard et al [3], published in this issue of The FEBS Journal, studied the role of wild-type BRAF during tumorigenesis, and particularly the mechanism controlling vascular permeability, a parameter which is deregulated during inflammation and cancer [4]. Increased permeability, with endothelial cells (ECs) as the main players, contributes to pathological features ranging from leukocyte or cancer cell transmigration (metastasis), fluid leakage (edema), poor drug delivery, to tumor angiogenesis [4].…”