Role of guanine nucleotide exchange factor P-Rex-2b in sphingosine 1-phosphate-induced Rac1 activation and cell migration in endothelial cells.
Endothelial cell (EC) migration has an important role in angiogenesis. Sphingosine-1 phosphate (S1P) stimulates EC migration via activation of Gi proteins. In this study, we characterized a mouse guanine nucleotide exchange factor (GEF) P-Rex2b for its regulation by Gbetagamma and PI3K and its role in S1P-induced Rac1 activation and cell migration in ECs. We found that co-expression of Gbetagamma or an active form of PI3K (PI3K(AC)) with P-Rex2b increased the SRE. Luciferase (SRE.L) reporter gene activity that can be stimulated by the Rho family of small GTPases including Rac1. Co-expression with P-Rex2b of Gbetagamma and PI3K(AC) or wild type PI3Kgamma that can be activated by Gbetagamma led to further increases in the reporter gene activity. Together with the finding that co-expression of Gbetagamma and/or PI3K(AC) increased the levels of active Rac1, we conclude that P-Rex2b is a Rac GEF that can be regulated by Gbetagamma and PI3K. Additionally, we demonstrated that Gbetagamma interacted with P-Rex2b, probably through P-Rex2b sequences at the PH domain and that the DEP and PDZ domains of P-Rex2b exerted an inhibitory effect on P-Rex2b's activity because their deletion increased the SER.L reporter gene activity. Furthermore, we found that P-Rex2b is involved in S1P-induced Rac1 activation and cell migration in ECs because siRNA-mediated suppression of P-Rex2b expression in ECs-diminished Rac1 activation and cell migration in response to S1P. Therefore, P-Rex2b is a physiologically significant Rac1 GEF that has an important role in the regulation of EC migration.
Li
Zhong
Z
Department of Genetics and Developmental Biology, University of Connecticut Health Center, Farmington, 06030, USA. zli@neuron.uchu.edu
Paik
Ji-Hye
JH
Paik
Jie-Hye
JH
Wang
Zhenglong
Z
Hla
Tim
T
Wu
Dianqing
D
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002465:Cell Movement / Q000502:physiology*; D017931:DNA Primers; D020690:GTPase-Activating Proteins / Q000502:physiology*; D020662:Guanine Nucleotide Exchange Factors; D006801:Humans; D047468:Immunoprecipitation; D008246:Lysophospholipids / Q000494:pharmacology*; D051379:Mice; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
D017931:DNA Primers; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D008246:Lysophospholipids; C490712:PREX2 protein, human; C060506:sphingosine 1-phosphate; D020830:rac1 GTP-Binding Protein; D013110:Sphingosine