SH3 domains direct cellular localization of signaling molecules.
In this study we describe the cellular distribution of the SH2 and SH3 domains of phospholipase C-gamma (PLC-gamma) and of the adaptor protein GRB2 following their microinjection into living rat embryo fibroblasts. Using immunofluorescence microscopy, we show that a truncated protein composed of the SH2 and SH3 domains of PLC-gamma was localized to the actin cytoskeleton. A similar localization pattern was observed when only the SH3 domain of PLC-gamma was microinjected. In contrast, a truncated protein composed of only the SH2 domains of PLC-gamma exhibited diffuse cytoplasmic distribution. Microinjected GRB2 protein was localized primarily to membrane ruffles, as was GRB2 protein containing SH2 loss-of-function point mutations. Hence, the localization of GRB2 to membrane ruffles does not require interaction with tyrosine-phosphorylated moieties. However, GRB2 proteins with SH3 loss-of-function point mutations exhibited diffuse cytoplasmic distribution. These results indicate that SH3 domains are responsible for the targeting of signaling molecules to specific subcellular locations.
Bar-Sagi
D
D
Cold Spring Harbor Laboratory, New York 11724.
Rotin
D
D
Batzer
A
A
Mandiyan
V
V
Schlessinger
J
J
D000199:Actins / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D002462:Cell Membrane / Q000737:chemistry; D002478:Cells, Cultured / Q000737:chemistry; D003593:Cytoplasm / Q000737:chemistry; D003599:Cytoskeleton / Q000378:metabolism*; D005347:Fibroblasts / Q000737:chemistry; D051380:GRB2 Adaptor Protein; D008856:Microscopy, Fluorescence; D011506:Proteins / Q000032:analysis*; D051381:Rats; D015398:Signal Transduction; D010738:Type C Phospholipases / Q000032:analysis*
D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495950:Grb2 protein, rat; D011506:Proteins; D010738:Type C Phospholipases