Intracellular signaling pathways involved in Gas6-Axl-mediated survival of endothelial cells

Abstract

Gas6 is a γ-carboxylated ligand for the receptor tyrosine kinase Axl. Gas6-Axl interactions can rescue endothelial cells from apoptosis, and this study examined the intracellular signaling mechanisms responsible for this phenomenon. Using flow cytometry, we first confirmed that Gas6 can abrogate apoptosis induced by serum starvation of primary cultures of human umbilical vein endothelial cells (HUVECs). This effect is mediated through phosphorylation of the serine-threonine kinase Akt, with maximal phosphorylation observed after 4 h of treatment with 100 ng/ml Gas6. Inhibition of Akt phosphorylation and abrogation of gas6-mediated survival of HUVECs by wortmannin implicated phosphatidylinositol 3-kinase as the mediator of Akt phosphorylation. Dominant negative Akt constructs largely abrogated the protective effect of Gas6 on HUVECs, underscoring the importance of Akt activation in Gas6-mediated survival. Several downstream regulators of this survival pathway were identified in HUVECs, namely, NF-κB as well as the antiapoptotic and proapoptotic proteins Bcl-2 and caspase 3, respectively. We showed that NF-κB is phosphorylated early after Gas6 treatment as evidenced by doublet formation on Western blotting. As well, the level of Bcl-2 protein increased, supporting the notion that the Bcl-2 antiapoptotic pathway is stimulated. The levels of expression of the caspase 3 activation products p12 and p20 decreased with Gas6 treatment, consistent with a reduction in proapoptotic caspase 3 activation. Taken together, these experiments provide new information about the mechanism underlying Gas6 protection from apoptosis in primary endothelial cell cultures.

Vimentin regulates EMT induction by Slug and oncogenic H-Ras and migration by governing Axl expression in breast cancer

“…Expression of Axl with its ligand Gas6 has been reported in many cancer types (Linger et al, 2008). Gas6-activated Axl signaling marshalls important downstream regulators of cell motility such as Rac and Akt (Allen et al, 2002;Hasanbasic et al, 2004) and regulates haptotaxis toward vitronectin in endothelial cells (Holland et al, 2005). Recent studies support a role for Axl as a regulator of glioma, breast, lung and pancreatic tumor cell migration (Shieh et al, 2005;Vajkoczy et al, 2006;Koorstra et al, 2009) and very recently Axl was identified as an important regulator of breast cancer metastasis and survival of patients (Vajkoczy et al, 2006;Gjerdrum et al, 2010).…”

Axl promotes cell invasion by inducing MMP-9 activity through activation of NF-κB and Brg-1

“…Previous studies indicated that Gas6 induces the phosphorylation of Axl in fibroblasts and endothelial cells, which contributes to some survival signaling, including the NF-kB transcription factor system (Demarchi et al, 2001;Hasanbasic et al, 2004). In NIH3T3 fibroblasts incubated in serum-deprived conditioned medium, the Gas6/Axl pathway upregulates Bcl-2 and Bcl-x(L) expression and activates NF-kB to suppress apoptosis.…”

Regulation of Microglial Phagocytosis and Inflammatory Gene Expression by Gas6 Acting on the Axl/Mer Family of Tyrosine Kinases

“…Gas6 is present at high levels in the brain throughout development and continues to be expressed in the adult (Prieto et al 1999). It has been reported that Gas6 has an anti-apoptotic effect on neurons (Goruppi et al 1996;Hasanbasic et al 2004), especially on protecting neurons from amyloid-beta-induced apoptosis (Yagami et al 2002). Furthermore, Gas6 may act as a neurotrophic factor for hippocampal (Funakoshi et al 2002) and cortical (Yagami et al 2003) neurons.…”