Covalent binding of guanine nucleotides to the CD3-gamma chain of the T cell receptor/CD3 complex.
In a search for proteins involved in signal transduction through the T cell receptor (TcR/CD3 complex), a recently developed highly efficient method for labeling of nucleotide binding proteins in permeabilized cells was applied. Here, we report that human CD3-gamma could be labeled by periodate-oxidized [alpha-32P] GTP (GTPoxi). In contrast to GTPoxi labeling of CD3-zeta (Peter, M. E., Hall, C., Rühlmann, A., Sancho, J. and Terhorst, C., EMBO J. 1992. 11:933), GTP-specific labeling of CD3-gamma reached a maximum when nucleotides were added 60 min prior to the cross-linking reaction. As CD3-gamma did not contain a known consensus sequence for nucleotide binding and since labeling kinetics of CD3-gamma coincided with those of cytosolic GTP-binding proteins, labeling may have been caused by a GTP-binding protein. This putative protein was not T cell specific because labeling of CD3-gamma could also be achieved when expressed in the endoplasmic reticulum of Chinese hamster ovary (CHO) cells. In CHO cells, labeling by GTPoxi took place only when CD3-gamma was associated with CD3-epsilon, whereas labeling could not be established upon association of CD3-gamma with CD3-delta or TcR alpha. The observation that CD3-gamma was labeled without leaving the endoplasmic reticulum led to the hypothesis that the association of CD3-gamma with a GTP-binding protein might be involved in an early step of the TcR/CD3 complex formation or transport.
Peter
M E
ME
Department of Medicine, Beth Israel Hospital, Harvard Medical School, Boston, MA 02215.
Wileman
T
T
Terhorst
C
C
D000818:Animals; D017252:CD3 Complex / Q000378:metabolism*; D016466:CHO Cells; D002460:Cell Line; D002478:Cells, Cultured; D006224:Cricetinae; D019204:GTP-Binding Proteins / Q000502:physiology; D006150:Guanine Nucleotides / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D017260:Receptor-CD3 Complex, Antigen, T-Cell / Q000378:metabolism*; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D016692:Receptors, Antigen, T-Cell, gamma-delta / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D013601:T-Lymphocytes / Q000378:metabolism; D014407:Tumor Cells, Cultured
D017252:CD3 Complex; D006150:Guanine Nucleotides; D017260:Receptor-CD3 Complex, Antigen, T-Cell; D011948:Receptors, Antigen, T-Cell; D016692:Receptors, Antigen, T-Cell, gamma-delta; C061836:antigen T cell receptor, epsilon chain; D019204:GTP-Binding Proteins