Fyn phosphorylates human MAP-2c on tyrosine 67.
The Src homology 3 (SH3) domain of Fyn binds to a conserved PXXP motif on microtubule-associated protein-2. Co-transfections into COS7 cells and in vitro kinase assays performed with Fyn and wild-type, or mutant MAP-2c, determined that Fyn phosphorylated MAP-2c on tyrosine 67. The phosphorylation generated a consensus sequence for the binding of the SH2 domain of Grb2 (pYSN). Pull-down assays with SH2-Grb2 from human fetal brain homogenates, and co-immunoprecipitation of Grb2 and MAP-2 confirmed the interaction in vivo, and demonstrated that MAP-2c is tyrosine-phosphorylated in human fetal brain. Filter overlay assays confirmed that the SH2 domain of Grb2 binds to human MAP-2c following incubation with active Fyn. Enzyme-linked immunosorbent assays confirmed the interaction between the SH2 domain of Grb2 and a tyrosine-phosphorylated MAP-2 peptide spanning the pY(67)SN motif. Thus, MAP-2c can directly recruit multiple signaling proteins important for central nervous system development.
Zamora-Leon
S Pilar
SP
Department of Pathology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Bresnick
Anne
A
Backer
Jonathan M
JM
Shafit-Zagardo
Bridget
B
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000378:metabolism; D020816:Amino Acid Motifs; D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers; D051380:GRB2 Adaptor Protein; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051076:Proto-Oncogene Proteins c-fyn; D014443:Tyrosine / Q000378:metabolism*; D018909:src Homology Domains; D019061:src-Family Kinases / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D017931:DNA Primers; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; C489788:MAP2 protein, human; D008869:Microtubule-Associated Proteins; D011518:Proto-Oncogene Proteins; D014443:Tyrosine; C494957:FYN protein, human; D051076:Proto-Oncogene Proteins c-fyn; D019061:src-Family Kinases