A general strategy for producing conditional alleles of Src-like tyrosine kinases.
The Src-like tyrosine kinases require membrane localization for transformation and probably for their normal role in signal transduction. We utilized this characteristic to prepare Src-like tyrosine kinases that can be readily activated with the rationally designed chemical inducer of dimerization FK1012. Dimerization of cytoplasmic Src-like tyrosine kinases was not sufficient for signaling, but their recruitment to the plasma membrane led to the rapid activation of transcription factors identical to those regulated by crosslinking the antigen receptor. Moreover, recruitment of activated Src-like kinases to the membrane replaced signaling by the T-lymphocyte antigen receptor complex, leading to the activation of both the Ras/protein kinase C and Ca2+/calcineurin pathways normally activated by antigen receptor signaling. Since these chemical inducers of dimerization are cell permeable, this approach permits the production of conditional alleles of any of the Src-like tyrosine kinases, thereby allowing a delineation of their developmental roles.
Spencer
D M
DM
Howard Hughes Medical Institute, Department of Developmental Biology, Stanford University Medical School, CA 94062, USA.
Graef
I
I
Austin
D J
DJ
Schreiber
S L
SL
Crabtree
G R
GR
D000469:Alkaline Phosphatase / Q000032:analysis; D000483:Alleles; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002451:Cell Compartmentation; D002478:Cells, Cultured; D018274:Electroporation; D004789:Enzyme Activation; D006801:Humans; D008213:Lymphocyte Activation / Q000235:genetics*; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D016296:Mutagenesis; D011493:Protein Kinase C / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D051076:Proto-Oncogene Proteins c-fyn; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics*; D016559:Tacrolimus / Q000031:analogs & derivatives / Q000494:pharmacology; D018909:src Homology Domains; D019061:src-Family Kinases / Q000235:genetics* / Q000378:metabolism*
C084260:FK 1012; D011518:Proto-Oncogene Proteins; D011948:Receptors, Antigen, T-Cell; D011993:Recombinant Fusion Proteins; C494957:FYN protein, human; D019860:Lymphocyte Specific Protein Tyrosine Kinase p56(lck); D051076:Proto-Oncogene Proteins c-fyn; D019061:src-Family Kinases; D011493:Protein Kinase C; D000469:Alkaline Phosphatase; D002118:Calcium; D016559:Tacrolimus