CD-3-mediated activation of MAP-2 kinase can be modified by ligation of the CD4 receptor. Evidence for tyrosine phosphorylation during activation of this kinase.
The CD4R has been shown to exert variable effects on T cell activation responses. Depending on the manner of ligation, the CD4R has been demonstrated to have positive as well as negative effects on the generation of [Ca2+]i flux by the CD3R. Coaggregation of CD3 with CD4 enhanced Ca2+ flux while their independent ligation and aggregation diminished this response. To further elucidate these paradoxical CD4 effects, we studied induction of a microtubule-associated protein 2 kinase (MAP-2K) activity during ligation of the CD3R. Lymphoid MAP-2K activation by CD3 is an evanescent event that is dependent on phosphorylation of 43-kDa MAP-2K via a pathway that involves protein kinase C. Coaggregation of CD4 and CD3 with cross-linking antibodies and avidin enhanced the CD3-mediated MAP-2K response almost twofold. In contrast, independent ligation and cross-linking of CD4 reduced the CD3-induced MAP-2K response by approximately 50%. An important requirement for this inhibitory effect was that CD4 be ligated before stimulation with anti-CD3. The negative effect of anti-CD4 mAb was specific as other mAb failed to simulate this event. The PMA-induced MAP-2K response was not inhibited by anti-CD4. Intact 32P-labeled Jurkat and normal human T cells demonstrated the appearance of a single 43-kDa tyrosine phosphoprotein during stimulation with PMA and anti-CD3. When these crude cellular extracts were extensively fractionated across DEAE- and hydrophobic columns, MAP-2K was resolved into two peaks of activity, each containing a single tyrosine phosphoprotein around 43 kDa. In addition to tyrosine-specific labeling, mitogenic stimulation of normal human T cells also induced threonine-specific labeling of MAP-2K. These results imply that activation of lymphoid MAP-2K is a dual process requiring at least two independent kinases for optimal activity. Inasmuch as CD3 activates protein kinase C and CD4 is associated with a tyrosine kinase, pp56lck, we suggest that their coaggregation may create the conditions whereby MAP-2K may be activated by dual phosphorylation. Independent aggregation of these receptors may lead to physical separation and breakdown of this interactive mechanism.
Nel
A E
AE
Department of Medicine, UCLA School of Medicine 90024-1680.
Pollack
S
S
Landreth
G
G
Ledbetter
J A
JA
Hultin
L
L
Williams
K
K
Katz
R
R
Akerley
B
B
D000328:Adult; D000945:Antigens, Differentiation, T-Lymphocyte / Q000276:immunology / Q000502:physiology*; D017252:CD3 Complex; D015704:CD4 Antigens / Q000276:immunology / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D004789:Enzyme Activation; D006801:Humans; D010766:Phosphorylation; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D011948:Receptors, Antigen, T-Cell / Q000276:immunology / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014443:Tyrosine / Q000378:metabolism*
D000945:Antigens, Differentiation, T-Lymphocyte; D017252:CD3 Complex; D015704:CD4 Antigens; D011948:Receptors, Antigen, T-Cell; D014443:Tyrosine; D011494:Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium