structural and Functional Studies of the T Cell Antigen Receptor 
The TCR activates a second signal transduction 
pathway that involves an enzyme with tyrosine 
kinase activity. The importance of the kinase 
pathway and its relationship to the inositol phos- 
pholipid pathway are being clarified by recent 
studies. We have found that stimulation of the 
TCR induces the tyrosine phosphorylation of an 
isozyme of phospholipase C, an enzyme that cata- 
lyzes the hydrolysis of PIP2. These studies, to- 
gether with additional studies using tyrosine ki- 
nase inhibitors, suggest that activation of a 
tyrosine kinase that is regulated by the TCR is 
required for activation of the phosphatidyl inosi- 
tol pathway. 
The mechanism by which the TCR couples to 
intracellular signaling pathways is largely unde- 
fined, as are many of the components of the sig- 
naling pathways themselves. To define and char- 
acterize the molecular basis by which the TCR 
regulates these pathways, we are using a somatic 
cell genetic approach. We have isolated a num- 
ber of mutants derived from T cell leukemic lines 
that are defective in TCR-mediated activation of 
the inositol phospholipid pathway. Unlike the pa- 
rental cells, none of these mutants produce lym- 
phokines in response to TCR stimulation. These 
mutants define four distinct gene products other 
than the Ti chains that are required for the func- 
tional activation of the inositol phospholipid 
pathway. 
In one mutant, the activation of the tyrosine 
kinase pathway is still intact. The defect in an- 
other of these mutants can be attributed to the 
absence of CD45, a cell surface protein with tyro- 
sine phosphatase activity. The absence of CD45 
prevents the TCR from activating the tyrosine ki- 
nase or phosphatidyl inositol pathway. This sug- 
gests a complex autoregulatory system, which we 
are intensively studying. In the remaining two 
mutants, biochemical studies have comple- 
mented our genetic approach and have suggested 
potential defects. We previously identified two 
molecules that associate with the TCR in a ligand- 
induced manner. In these two mutants, these mol- 
ecules fail to associate with the TCR. Further stud- 
ies of the basis for the defective signal 
transduction observed in these mutants are in pro- 
gress. Thus these mutants are proving to be valu- 
able tools with which to dissect the complexities 
of the signal transduction pathways and their re- 
lationships to cellular responses. 
An alternative approach toward understanding 
which signal transduction pathways can lead to 
cellular responses is to express other heterolo- 
gous receptors with well-defined signal transduc- 
tion functions in T cells and examine their abili- 
ties to activate typical T cell responses. For 
example, the human muscarinic acetylcholine 
receptor subtype 1 (HMl) activates the inositol 
phospholipid pathway in cardiac, smooth mus- 
cle, and neuronal cells. We have expressed HMl 
in a T cell line and find that, when stimulated, it 
can activate the inositol phospholipid pathway 
but not a tyrosine kinase pathway. Moreover, 
HMl stimulation results in lymphokine produc- 
tion and other typical results associated with TCR 
stimulation. These results suggest that the inosi- 
tol phospholipid pathway, a signal transduction 
pathway common to both the TCR and HMl , does 
regulate T cell responses. How this pathway 
serves to regulate later cellular responses is being 
addressed in ongoing studies. 
T cell activation is a complex process that is 
regulated by cell surface molecules. Investiga- 
tion of the molecules and events involved in the 
activation of T cells should lead to a more com- 
plete understanding of T cell biology and a more 
rational approach to the manipulation of the im- 
mune system. Moreover, through the study of the 
activation of T cells, it is likely that insight into 
other biological systems involving cell prolifera- 
tion and differentiation will emerge. 
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