The guanine nucleotide exchange factor for Rac P-Rex1

Researchers:

Sylvia Thelen, Scientist

Status: In progress

The small GTPases of the Rho family, Rac and Cdc42, are critical for rapid rearrangements of the actin cytoskeleton observed during filopodia and lamellipodia formation in migrating cells. The GTPases act as switches and are either ‘on’ in their GTP bound form or ‘off’ when loaded with GDP. Activation of the GTPases is catalyzed by specific GTP exchange factors (GEF). The phosphatidylinositol 3,4,5-trisphosphate (PIP₃)-dependent exchanger 1 (P-Rex1) is assumed to be involved in G-protein coupled receptor (GPCR)-mediated Rac activation. P-Rex1 activity is stimulated by the PI 3-kinase product PIP3 and by the βγ subunits of heterotrimeric G-proteins, which are released upon activation of GPCRs. Consistent with the activation by these cofactors and their cellular localization following stimulation of the cells, P-Rex1 is recruited to the plasma membrane. Overexpression of P-Rex1 or its suppression by siRNA markedly alters chemokine-stimulated migratory capacity of myeloid leukocytes, consistent with the assumption that GEF is required for efficient chemotaxis. P-Rex1 becomes phosphorylated at multiple sites following cell activation and the modification appears to contribute to its subcellular localization. The project aims for the molecular characterization of different P-Rex1 domains and their role in chemokine receptor-mediated signal transduction. To this end, various P-Rex1 mutants will be transduced into hematopoietic precursor cells, which are derived from animals with a genetic deletion of P-Rex1. The cells can be differentiated later into neutrophils and monocytes and tested for the activity of the respective P-Rex1 mutants. To further investigate the expression of P-Rex1 in different tissues and to better reveal its subcellular localization, we generated monoclonal antibodies suitable for immunofluorescence analysis and immunohistochemistry. Staining of normal epithelium and invasive cancer from human esophagus reveals a marked upregulated expression of P-Rex1 in the tumor. The data suggest a potential role of P-Rex1 for tumor cell migration and invasion.

Welch, H.C., A.M.Condliffe, L.J.Milne, G.J.Ferguson, K.Hill, L.M.Webb, K.Okkenhaug, W.J.Coadwell, S.R.Andrews, M.Thelen, G.E.Jones, P.T.Hawkins, and L.R.Stephens. 2005. P-rex1 regulates neutrophil function. Curr.Biol. 15:1867-1873.
Barber,M.A., S.Donald, S.Thelen, K.E.Anderson, M.Thelen, and H.C.Welch. 2007. Membrane translocation of P-Rex1 is mediated by G protein beta gamma subunits and phosphoinositide 3-kinase. J.Biol.Chem. 282:29967-29976.