The function of the signaling protein Ras guanine releasing protein 4 (RasGRP4) in human mast cells

Mast cells have been implicated in the pathogenesis of both atopic and non-atopic asthma. Rasguanine nucleotide-releasing protein 4 (RasGRP4) is a mast cell-restricted guanine nucleotideexchange factor and diacylglycerol (DAG)/ phorbol ester receptor whose function has not beendeduced. RT-PCR analysis of 40 asthmatic patients and 40 non-asthmatic controls demonstrated ahigher hRasGRP4 mRNA expression in a subgroup of the asthmatics. A RasGRP4-defective variantof the human mast cell line HMC-1 was used to create stable clones expressing green fluorescentprotein-labeled human RasGRP4 for monitoring the movement of this signaling protein inside mastcells before and after exposure to phorbol-12-myristate 13-acetate (PMA) and for evaluating theprotein’s ability to control the development, phenotype, and function of mast cells.Transcript-profiling approaches revealed hRasGRP4 constitutively regulates the expression ofnumerous genes in the HMC-1 cell line. For example, expression of hRasGRP4 in HMC-1 cellssubstantially decreased GATA-1 levels without altering GATA-2 levels, suggesting that hRasGRP4regulates mast cell commitment of multipotential progenitors in part by controlling the intracellularlevels of at least one lineage-dependent transcription factor for hematopoietic cells. hRasGRP4resided primarily in the cytosol before HMC-1 cells were stimulated with PMA. After exposure toPMA, hRasGRP4 translocated to the inner leaflet of the cell’s plasma membrane and then toperinuclear and Golgi compartments. Extracellular signal-regulated kinases 1 and 2 were activatedduring this translocation process, and the PMA-treated cells transiently increased their expression ofthe transcripts encoding the interleukin 13 receptor IL-13Rá2 and numerous other proteins.The accumulated data in our mast cell model suggest hRasGRP4 translocates to various intracellularcompartments via its DAG/PMA-binding domain to regulate those signaling pathways that allow mastcells to respond quickly to changes in their tissue microenvironments