The nitric oxide/cGMP pathway regulates neutrophil adhesion through activation of rap GTPases

• Main Author: Deevi, Ravi Kiran
• Published: Queen's University Belfast 2008
• Subjects: 612
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.492153

We found that the pro-inflammatory cytokine TNF-ex. promoted adhesion of human neutrophils to fibrinogen. However, TNF- ex. activated Rap GTPases in adherent but not suspended neutrophils. Thus, signalling through ~2 integrins is required to initiate activation of Rap GTPases. We have clearly shown that activation of Rap1 and Rap2 by ~2 integrins requires production of NO and cGMP, as well as activation of cGMPdependent serine/threonine kinases (cGKI). We also found that activation of iNOS by ~2 integrins required Src tyrosine kinases, Protein Kinase C, Phosphoinostide 3-Kinase, and calcium signalling. We also discovered that Rap2 might be negatively regulated via an unknown pathway and independently from production ofNO. Similarly, we found that the formyl peptide fMLP activates Rap GTPases and adhesion in human neutrophils and differentiated PLB-985 cells, through production of NO and activation of cGKI. Furthermore, we discovered that VASP, a cytoskeletal protein is phosphorylated on serine 239 in response to tMLP or a cGMP analogue in leucocytes. We further discovered that VASP and polymerization of actin is essential for activation of Rap GTPases and adhesion of leucocytes to fibrinogen. Indeed, silencing VASP in differentiated PLB-985 cells abolished cGMP-induced activation of Rap GTPases and adhesion. In fact, we demonstrated that VASP is a negative regulator of Rap GTPases in resting cells, since silencing VASP in differentiated PLB-985 cells resulted in augmented basal Rap GTPases activities and adhesion. Interestingly, silencing VASP in differentiated PLB-985 cells also induced redistribution of C3G (a guanine nucleotide exchange factor for Rap GTPases) and Rapl to the plasma membrane. In summary, we propose that phosphorylation of VASP by cGKI which occurs in response to fMLP is responsible for polymerization of actin, and the associated augmented rigidity of the cytoskeleton confers the physical force required to redistribute C3G to the plasma membrane where Rap GTPases are activated. Supplied by The British Library - 'The world's knowledge'