FIGURE 1.

BCR-induced plasma membrane targeting of RasGRP1 is mediated by its BHC. A, domain structure of RasGRP1, the sequence of the PT domain and the boundaries of the PT-N and PT-C segments, and the BHC and leucine zipper (LZ). Numbering corresponds to the sequence of full-length murine RasGRP1. The sequence changes for mutants are shown below the PT sequence. B and D, DT40 cells expressing GFP-tagged RasGRP1 (RG1) or the indicated derivatives were untreated (nil) or treated with α-IgM for 10 min and then imaged, as described under “Experimental Procedures.” The single cells shown in this and subsequent figures are representative of the majority of cells in the population, with the exception of those expressing PT-C that were chosen to represent the subset of cells with detectable plasma membrane localization of PT-C (19% of the cell population for nil, and 20% for α-IgM-treated). Quantifications of signal at plasma membrane versus cytoplasm are in Table 1. Cells shown have diameters of 12 ± 1 μm unless otherwise indicated by the presence of a scale bar. Scale bars, 5 μm. C, sequence comparison of the PT-N segment of vertebrate RasGRP1s. Conserved amino acids are boxed (R = K and D = E for conservation). Basic amino acids are blue, and large hydrophobic amino acids are green.