Specific interaction of the CD45 protein-tyrosine phosphatase with tyrosine-phosphorylated CD3 zeta chain.

Abstract

The CD45 transmembrane protein-tyrosine phosphatase (PTPase, EC 3.1.3.48) plays an essential role in T-cell activation by activating the Lck and/or Fyn proteintyrine kinases. However, numerous experiments have indicated that CD45 may have both mulatory and inhibitory roles in T-cell activation. Thus, it is unlikely that the two kinases are the sole substrates of the CD45 PTPase. Furthermore, the complex regulation of the alternative splicing of the extracellular domain in various leukocyte lineages also suggests additional roles for the CD45 PTPase. To identify such functions, it is necessary to identify physiologically relevant substrates of the CD45 PTPase other than the two protein-tyrosine kinases. To this end, we searched for high-affinity substrates of the CD45 VPTase among the tyrosine-phosphorylated T-cell proteins by using purified glutathione S-transferase-CD45 fusion molecules. The enzymatically inactive CD45 C828S mutant protein, in which the cysteine residue at the catalytic center was changed to a serine residue, bound tightly to the phosphorylated CD3 gchailn. This binding was specific to CD45 PTPase, as neither the leukocyte common antigen-related molecule (LAR) PTPase nor the CD45-LAR hybrid PTPases bound the phosphorylated CD3 S chain. Furthermore, phosphorylated CD3 C chain was preferentially dephosphorylated by the wild-type CD45 PTPase under conditions that did not significantly dephosphorylate other cellular proteins. Thus, the phosphorylated CD3 S chain is a specific and high-affinity substrate of the CD45 PTPase. These results suggest that CD45 is involved in the termination of the T-cell response via dephosphorylation of CD3 S chain.

Phosphatase CD45 Both Positively and Negatively Regulates T Cell Receptor Phosphorylation in Reconstituted Membrane Protein Clusters

“…However, CD3 phosphorylation levels were sharply decreased in the presence of only 50 -100 m Ϫ2 CD45 molecules, which is significantly lower than the physiological CD45 density in T cell membranes (ϳ1000 m Ϫ2 ) (34). This trend is in good agreement with previous measurements in solution phase or on liposomes (6,24). CD3 phosphorylation in the absence of CD45 appeared to saturate at lower Lck density (ϳ100 m Ϫ2 ) in homogeneous membranes, whereas that in His 10 -protein clusters increased with Lck density (100 -400 m Ϫ2 ), suggesting a slower reaction rate in the clusters (Fig.…”

“…TCR phosphorylation and subsequent signaling also require a highly expressed transmembrane phosphatase, CD45 (3)(4)(5). However, TCR is also a substrate of CD45 (6). Thus, CD45 has dual positive and negative roles in the induction of TCR phosphorylation (7).…”

Cell‐type specificity of anti‐CD45 autoantibodies in systemic lupus erythematosus

“…In addition to its crucial role in the initiation of signaling, CD45 may be involved in the termination of T cell responses (20). Depending on the nature of the activation signals, the experimental conditions of the assays, and the state of lymphocyte activation, different anti-CD45 monoclonal antibodies (MAb) profoundly influence cell function.…”

Molecular cloning and characterization of Byp, a murine receptor‐type tyrosine phosphatase similar to human DEP‐1

“…PTPs with the Cys to Ser mutation have been reported to associate stably with their substrates. For example, the Cys to Ser mutants of 3CHl34 and CD45 well associate with ~44~~~~ [32] and the CD31 chain [33], respectively. Therefore, the Cys to Ser mutant could help identify the substrate of each PTP.…”