Role of Epac1, an Exchange Factor for Rap GTPases, in Endothelial Microtubule Dynamics and Barrier Function

Abstract

Rap1GTPase activation by its cAMP responsive nucleotide exchange factor Epac present in endothelial cells increases endothelial cell barrier function with an associated increase in cortical actin. Here, Epac1 was shown to be responsible for these actin changes and to colocalize with microtubules in human umbilical vein endothelial cells. Importantly, Epac activation with a cAMP analogue, 8-pCPT-2O-Me-cAMP resulted in a net increase in the length of microtubules. This did not require cell-cell interactions or Rap GTPase activation, and it was attributed to microtubule growth as assessed by time-lapse microscopy of human umbilical vein endothelial cell expressing fluorophore-linked microtubule plus-end marker end-binding protein 3. An intact microtubule network was required for Epac-mediated changes in cortical actin and barrier enhancement, but it was not required for Rap activation. Finally, Epac activation reversed microtubuledependent increases in vascular permeability induced by tumor necrosis factor-␣ and transforming growth factor-␤. Thus, Epac can directly promote microtubule growth in endothelial cells. This, together with Rap activation leads to an increase in cortical actin, which has functional significance for vascular permeability.

Single-cell analyses of transcriptional heterogeneity during drug tolerance transition in cancer cells by RNA sequencing

“…Two of them resided in genes that were previously reported to be involved in paclitaxel resistance. One was a missense mutation located in RAPGEF4 that encodes a protein involved in microtubule polymerization and organization (33,34). The other was found in the 3′ UTR region of AMOTL1.…”

“…Three other RNA variants unique to the drug-tolerant single cells were found in genes that were involved in microtubule organization and stabilization during mitosis: RAPGEF4, NUDCD3, and KIAA1671 (Table 1). RAPGEF4 (Rap guanine nucleotide exchange factor 4) was previously shown to interact with protein complexes that were involved in microtubule polymerization and organization (33,34). RAPGEF4 protein is also known as exchange protein directly activated by cAMP 2 (EPAC2) and is one of the binding partners of MAP1A (microtubule-associated protein 1A) (33).…”

The Epac-Rap1 Signaling Pathway Controls cAMP-mediated Exocytosis of Weibel-Palade Bodies in Endothelial Cells

“…and thereby abolishes VWF secretion (12). Epac, the exchange protein activated by cAMP for the small GTPases Rap1 and Rap2, is involved in regulation of endothelial barrier function (27)(28)(29) and endothelial cell adhesion (30) but also regulated secretion of insulin in pancreatic beta-cells (31). Endothelial cells selectively express Epac1 but not Epac2 (28,29).…”

“…Epac, the exchange protein activated by cAMP for the small GTPases Rap1 and Rap2, is involved in regulation of endothelial barrier function (27)(28)(29) and endothelial cell adhesion (30) but also regulated secretion of insulin in pancreatic beta-cells (31). Endothelial cells selectively express Epac1 but not Epac2 (28,29). It has been shown previously that the Epac-specific cAMP analog 8-pCPT-2Ј-O-Me-cAMP promotes exocytosis of WPBs (9,32).…”

The role of cytoskeleton in the regulation of vascular endothelial barrier function

“…Rap1 GEF Epac is cAMP-responsive protein, which provides PKA-independent regulation of cytoskeleton organization. Epac localizes with microtubules; its specific activation with cAMP analogue o-Me-cAMP results in microtubule elongation, and, more importantly, reverses microtubule-dependent increases in vascular permeability induced by TNFα and TGFβ [Sehrawat et al, 2008]. Surprisingly, this effect of Epac was found to be Rap1-independent.…”