Submitted January 10, 2007
Accepted February 26, 2007
Roles of RabGEF1/Rabex-5 domains in regulating Fc
RI surface expression and FcRI-
dependent responses in mast cells
Janet Kalesnikoff, Eon J Rios, Ching-Cheng Chen, M. Alejandro Barbieri, Mindy Tsai, See-Ying Tam, and Stephen J. Galli*
Department of Pathology, Stanford University School of Medicine, Stanford, CA, United States
Department of Biological Sciences, Florida International University, College of Arts and Sciences, Miami, FL, United States
* Corresponding author; email: sgalli{at}stanford.edu.
RabGEF1/Rabex-5, a guanine nucleotide exchange factor (GEF) for the endocytic pathway regulator, Rab5, contains a Vps9 domain, an A20-like zinc finger (ZnF) domain and a coiled coil domain. To investigate the importance of these domains in regulating receptor internalization and cell activation, we lentivirally delivered RabGEF1 mutants into RabGEF1-deficient (-/-) mast cells and examined Fc
RI-dependent responses. Wild-type RabGEF1 expression corrected phenotypic abnormalities in -/- mast cells, including decreased basal FcRI expression, slowed FcRI internalization, elevated IgE+Ag-induced degranulation and IL-6 production, and the decreased ability of -/- cytosol to support endosome fusion. We showed that RabGEF1's ZnF domain has ubiquitin ligase activity. Moreover, the coiled coil domain of RabGEF1 is required for Rabaptin-5 binding and for maintaining basal levels of Rabaptin-5 and surface FcRI. However, mutants lacking either of these domains normalized phenotypic abnormalities in IgE+antigen-activated -/- mast cells. By contrast, correction of these -/- phenotypes required a functional Vps9 domain. Thus, FcRI-mediated mast cell functional activation is dependent on RabGEF1's GEF activity.
