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Cutting Edge: The CD45 Tyrosine Phosphatase Is an Inhibitor of Lck Activity in Thymocytes

Laboratory of Immune Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892

	Abstract

Top Abstract Introduction Materials and Methods Results and Discussion References

 
A widely accepted model for regulation of the Lck tyrosine kinase is that it is activated by CD45-mediated dephosphorylation of its COOH-terminal negative regulatory tyrosine (Tyr⁵⁰⁵). Previous work from our laboratory, however, found that despite hyperphosphorylation of Tyr⁵⁰⁵, the activity of Lck from CD45^- T cell lines was actually increased due to hyperphosphorylation of the positive regulatory tyrosine, residue 394. To avoid potential complications introduced by transformed cells, in this study we have characterized the effect of CD45 on Lck activity in normal cells. Lck in thymocytes from CD45^-/- mice was hyperphosphorylated on tyrosine residues. Importantly, and in disagreement with the model that CD45 only activates Lck in vivo, the kinase activity of Lck from cells lacking CD45 was substantially increased. These results support a model in which CD45 dephosphorylates both Tyr⁵⁰⁵ and Tyr³⁹⁴, the net effect in normal thymocytes being a decrease in enzymatic activity.

	Introduction

Top Abstract Introduction Materials and Methods Results and Discussion References

 
Activation of protein tyrosine kinases (PTKs)² is an early and critical step in the signal transduction cascade originating from the TCR, and their inhibition is sufficient to prevent induction of almost all T cell responses 1, 2, 3 . Among the PTKs that have been implicated in this process are two members of the src family of tyrosine kinases: Fyn and Lck. The activity of src family kinases is regulated by the phosphorylation and dephosphorylation of two key tyrosine residues. One, in the COOH-terminal region (Tyr⁵⁰⁵ for murine Lck), is the major site of negative regulation 4, 5 . The other, in the kinase domain, enhances kinase activity when phophorylated. This tyrosine (residue 394 for murine Lck) is the major site of autophosphorylation 6 . The kinase activity of Lck and Fyn is therefore subject to regulation by enzymes that dephosphorylate these crucial tyrosines. The most abundant of these protein tyrosine phosphatases in hemopoietic cells is the transmembrane molecule CD45. Indeed, CD45^- T cells have profound defects in TCR signaling 7, 8, 9 , and reconstitution of these cells with the catalytically active and membrane-associated CD45 phosphatase domain is sufficient to restore normal signaling 10, 11 . Since the COOH-terminal tyrosines of Lck and Fyn are good substrates for CD45 in vitro 12, 13 and phosphorylation of these residues is inhibitory for kinase activity, CD45 has been considered as an activator of src family kinases. An analysis of the published data, however, suggests that the relationship between CD45 and src family kinase activity is not so simple. While there are many reports showing that in CD45^- T cell lines Lck is hyperphosphorylated on the COOH-terminal tyrosine 14, 15, 16, 17 , there is a lack of consistency between the studies on the influence of this hyperphosphorylation on PTK activity. Work from our laboratory using CD45^- T cell lines found that Lck kinase activity was increased despite hyperphosphorylation of the COOH-terminal tyrosine residues 18 and that this was due to hyperphosphorylation of Tyr³⁹⁴ 19 .

It has been suggested that some of the differences between studies on the role of CD45 on src family kinase activity may be due to the nature of the cells, which are typically retrovirally transformed and therefore potentially subject to the effects of viral oncogenes 20 . To address this, we now analyze the enzymatic activity of Lck in thymocytes from CD45-deficient mice 21 . Although Lck is hyperphosphorylated on tyrosine residues, its kinase activity is substantially increased in the absence of CD45, demonstrating that in normal lymphoid cells CD45 inhibits the activity of this src family kinase.

	Materials and Methods

Top Abstract Introduction Materials and Methods Results and Discussion References

 
Cells and Abs

Mice with disruption of CD45 exon 9 on the C57BL/6J background 21 and normal C57BL/6J mice were purchased from The Jackson Laboratory (Bar Harbor, ME). Individual thymi were harvested, and single-cell suspensions were prepared in RPMI 1640 (Biofluids, Gaithersburg, MD) supplemented with 10% FCS. Cells were washed with PBS, counted, and used for flow cytometry analysis or biochemical studies. Anti-CD4-PE (RM4-5) and anti-CD8-FITC (53-6.7) were purchased from PharMingen (San Diego, CA). Anti-CD45 (M1/89) was used as culture supernatant and detected with FITC-goat anti-rat (Jackson ImmunoResearch, West Grove, PA). Anti-Lck mAb 3A5 bound to agarose beads and anti-phosphotyrosine Ab PY-99 were obtained from Santa Cruz Biotechnology (Santa Cruz, CA). Lck antiserum 688 was kindly provided by Larry Samelson, National Institutes of Health, Bethesda, MD.

Immunoprecipitation and immunoblotting

Cells were washed in cold PBS and lysed in buffer containing 50 mM Tris, 300 mM NaCl, 0.5% Triton X-100, leupeptin, aprotinin, and phosphatase inhibitors 2 mM sodium o-vanadate, 0.4 mM EDTA, 10 mM sodium fluoride, and 10 mM sodium pyrophosphate (lysis buffer). Immunoprecipitation was performed on postnuclear fractions for 2 h at 4°C with protein A-Sepharose beads (Pierce, Rockford, IL) precoated with the indicated Abs. The precipitated proteins or total cell lysates were separated on reducing 8% SDS-PAGE, transferred to nitrocellulose, and immunoblotted with the indicated Abs. Immunoblots were developed by ¹²⁵I-labeled protein A and analyzed with a PhosphorImager (Molecular Dynamics, Sunnyvale, CA).

In vitro tyrosine kinase assay

PTK activity was assayed using phosphorylation of the peptide KVEKIGEGTYGVVKK from p34^cdc2 residues 6–20 (Pierce) as described 22 . Briefly, washed cells were lysed in lysis buffer, and postnuclear fractions were immunoprecipitated with Lck antiserum 688. After extensive washing, the immunoprecipitates were incubated with 21 µl of a reaction mixture containing the cdc2 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 peptide substrate and 10 µCi of [-³²P]ATP (ICN) at 30°C for 10 min. Blank values were obtained for each sample in the absence of substrate. Reactions were stopped by adding 10 µl of 10% acetic acid, the samples were centrifuged in an Eppendorf microfuge (Hamburg, Germany) for 30 s, and 25 µl of supernatant from each sample were spotted on a SpinZyme basic separation unit (Pierce). After two washes with 75 mM phosphoric acid, bound radioactivity was counted with a ß-scintillation counter. At the same time, the immunocomplexes used for the kinase reaction as well as for the blank reaction (with no peptide) were washed three times, resuspended in sample buffer, and separated on reducing 8% SDS-PAGE to analyze Lck autophosphorylation. Gels were transferred to nitrocellulose, and radioactivity was measured by PhosphorImager (Molecular Dynamics).

	Results and Discussion

Top Abstract Introduction Materials and Methods Results and Discussion References

 
Disruption of CD45 exon 9 in mice leads to the complete absence of CD45 expression in hemopoietic cells 21 . Cytofluorometric analysis of these animals confirmed the absence of CD45 on thymocytes as well as a previously reported defect in the transition from CD4⁺CD8⁺ to the more mature CD4⁺CD8^- and CD4^-CD8⁺ stages of differentiation (Fig. 1). To characterize Lck expression and activity, equal amounts of total protein lysates from the CD45^-/- and CD45^+/+ thymocytes were separated on SDS-PAGE and blotted with an Lck-specific antiserum. Thymocytes from these mice expressed Lck at comparable levels (Fig. 2A). The tyrosine phosphorylation status of Lck was determined by immunoprecipitating it from equal amounts of cell lysates and immunoblotting with an anti-phosphotyrosine Ab. Consistent with data obtained with CD45^- cell lines and a previous analysis of these mice 21 , Lck isolated from CD45^-/- thymocytes had a much higher degree of tyrosine phosphorylation than Lck obtained from CD45^+/+ thymocytes (Fig. 2B).

View larger version (40K): [in this window] [in a new window]   FIGURE 1. Flow cytometry analysis of CD45-/- and CD45+/+ thymocytes. Thymocytes from CD45-/- and CD45+/+ mice were stained for CD4 and CD8 (top) or CD45 (bottom) and analyzed with a FACScan.

View larger version (22K): [in this window] [in a new window]   FIGURE 2. Lck in CD45-/- thymocytes is hyperphosphorylated on tyrosine residues. A, Lysates of CD45-/- and CD45+/+ thymocytes were separated on 8% SDS-PAGE and immunoblotted with anti-Lck. B, Lck was immunoprecipitated from CD45-/- and CD45+/+ thymocytes using Ab 3A5 covalently coupled to agarose beads. The material was separated by 8% SDS-PAGE and immunoblotted with anti-phosphotyrosine Ab PY-99. The lane labeled Ab alone is a sham immunoprecipitation with no lysate.

View larger version (21K): [in this window] [in a new window]   FIGURE 3. Measurement of Lck autophosphorylation activity. A, Lck was immunoprecipitated from CD45-/- and CD45+/+ thymocytes and the material subjected to immune complex kinase assay (top) or immunoblot with anti-Lck antiserum (bottom). The reactants were separated by 8% SDS-PAGE. B, PhosphorImager analysis of three independent autophosphorylation experiments using three pairs of CD45-/- and CD45+/+ mice. The geometric mean and the SEM (x/÷) are shown.

View larger version (14K): [in this window] [in a new window]   FIGURE 4. Assessment of Lck activity by measuring phosphorylation of an independent substrate. A, Lck was immunoprecipitated from CD45-/- and CD45+/+ thymocytes and was used to phosphorylate p34cdc2 (6–20). B, Results of 5 independent experiments in which the phosphorylation of p34cdc2 (6–20) by Lck was measured. The geometric mean and the SEM (x/÷) are shown.

Despite the commonly held assumption that CD45 activates Lck in vivo 25, 26 , based largely on the analyses of Tyr⁵⁰⁵ phosphorylation, there is surprisingly little direct evidence for this model, and in fact quite a bit of data indicating that the relationship is not so straightforward. In favor of a direct positive effect of CD45 on the activation of Lck is a report in which Lck and Fyn from a CD45^- CD8⁺ IL-2-dependent T cell clone were hyperphosphorylated in the COOH-terminal region and had reduced kinase activity 14 . In another report using a CD45^- human leukemic T cell line, the Lck coprecipitated with anti-CD4 was reported to have reduced kinase activity 27 . However, in the same CD45^- cells, the activity of Lck immunoprecipitated from whole cell lysates was clearly elevated. Moreover, another analysis of CD45^- variants of the T cell line HPB-ALL found an increase in the kinase activity of CD4-associated Lck 28 . Cross-linking CD4 further enhanced Lck activity in these CD45^- cells as well as in CD45^- SAKRTLS cells 29 , demonstrating that CD45 is not required for induction of Lck activity. Other data in the literature also challenge the simple model that CD45 as a pure activator of src family kinases. For example, coclustering of CD45 with CD4 inhibits both Lck phosphorylation and increases in kinase activity induced by CD4 engagement 29 , suggesting that CD45 dephosphorylation of Lck decreases rather than increases Lck activity. In agreement with this is the finding that inhibition of tyrosine phosphatase activity by pervanadate is associated with cellular hyperphosphorylation and activation of Lck and Fyn and that these events depend on inhibition of CD45 since they do not occur in CD45^- cells 30, 31 . Furthermore, in one set of CD45^- Jurkat T cells, the baseline activities of Lck and Fyn have been shown to be elevated 31 . We have demonstrated in three independent T cell lines that in the absence of CD45 src family kinases are hyperphosphorylated, predominantly on Tyr⁵⁰⁵ 18 . Despite this, Lck and Fyn activity were substantially elevated, and this activity was reduced to baseline levels by in vitro exposure of the hyperphosphorylated Lck to CD45. Phosphopeptide mapping and the characterization of Lck mutants with substitutions of critical tyrosine residues with phenylalanine demonstrated that loss of CD45 loss resulted in increased phosphorylation not only of Tyr⁵⁰⁵ but also of Tyr³⁹⁴ and that the latter was responsible for the increased kinase activity 19 . These results offer one reason for the apparent discrepancies between some of the studies, namely, that depending on the balance of phosphorylation between Tyr³⁹⁴ and Tyr⁵⁰⁵ one might obtain either inhibition or activation of kinase activity.

A previous study with a different CD45 knockout strain reported that the activity of the src family kinases Hck and Lyn were in fact elevated, although the activity of Lck in these mice was not determined 32 . Another study with the mice used in the present report demonstrated that Lck and Fyn were hyperphosphorylated on tyrosine residues and used Lck precipitation by a peptide containing phosphorylated Tyr⁵⁰⁵ that cannot bind the "closed form" of the molecule (i.e., phosphorylated at Tyr⁵⁰⁵) to suggest that there was more kinase in the inactive form 33 . This result is not surprising, as loss of CD45 is always associated with increased phosphorylation of Tyr⁵⁰⁵. However, since loss of CD45 also results in increased phosphorylation of Tyr³⁹⁴ 19 , this does not mean that the net kinase activity in the population of Lck molecules will in fact be decreased. No direct assay of overall Lck kinase activity was performed in that study 33 . In the current report, we have directly quantitated the enzymatic function of Lck. In accord with our previous data with CD45^- T cell lines, despite tyrosine hyperphosphorylation, Lck from CD45^-/- thymocytes was 2–4 times more active than Lck from CD45^+/+ thymocytes. This is similar to the degree to which kinase activity is elevated in the "constitutively active" form of Lck in which Tyr⁵⁰⁵ is replaced with a Phe 19 . These findings, together with previous observations 18, 19, 30, 31 , support an alternative to the simple model that CD45 merely activates Lck by dephosphorylation of Tyr⁵⁰⁵ (Fig. 5). In this scheme, CD45 is able to dephosphorylate both Tyr⁵⁰⁵ and Tyr³⁹⁴, and the consequence of these activities in normal thymocytes is a decrease in Lck kinase activity. Therefore, one direct role for CD45 in normal T cell activation would be to reset Lck to its hypophosphorylated status at the end of the TCR signaling cascade, bringing the kinase back to its preactivation status. Together, these results indicate a more complex role for CD45 on src family kinase activity than is typically held and demonstrate that in normal nontransformed thymocytes CD45 down-regulates Lck activity.

View larger version (10K): [in this window] [in a new window]   FIGURE 5. Schematic of how CD45 influences Lck activity by dephosphorylating regulatory tyrosine residues. Lck can be phosphorylated at Tyr505 (left) or Tyr394 (right), or possibly both (not shown). CD45 dephosphorylates both tyrosine residues, which can either enhance or diminish Lck activity, depending on the relative abundance of Tyr505 vs Tyr394 phosphorylation. In thymocytes, the net effect is to diminish Lck activity. Lck in which Tyr394 has been substituted with a Phe is inactive, even when Tyr505 is substituted with a Phe, and therefore cannot be phosphorylated (19); this implies that this phosphorylation of Tyr394 is a prerequisite for Lck activity. The issue of whether there are Lck molecules phosphorylated on both Tyr394 and Tyr505, and if so what would the kinase activity of such a molecule be, has not been experimentally addressed.

	Acknowledgments

 
We thank Allan Weissman for critical review of the manuscript.

	Footnotes

 
¹ Address correspondence and reprint requests to Dr. Jonathan Ashwell, Laboratory of Immune Cell Biology, National Cancer Institute, National Institutes of Health, 9000 Rockville Pike, Building 10, Room 1B40, Bethesda, MD 20892-1152. E-mail address:

² Abbreviation used in this paper: PTKs, protein tyrosine kinases.

Received for publication November 4, 1998. Accepted for publication December 10, 1998.

	References

Top Abstract Introduction Materials and Methods Results and Discussion References

 

1. June, C. H., M. C. Fletcher, J. A. Ledbetter, L. E. Samelson. 1990. Increases in tyrosine phosphorylation are detectable before phospholipase C activation after T cell receptor stimulation. J. Immunol. 144:1591.[Abstract]
2. June, C. H., M. C. Fletcher, J. A. Ledbetter, G. L. Schieven, J. N. Siegel, A. F. Phillips, L. E. Samelson. 1990. Inhibition of tyrosine phosphorylation prevents T-cell receptor-mediated signal transduction. Proc. Natl. Acad. Sci. USA 87:7722.[Abstract/Free Full Text]
3. Hanke, J. H., J. P. Gardner, R. L. Dow, P. S. Changelian, W. H. Brissette, E. J. Weringer, B. A. Pollok, P. A. Connelly. 1996. Discovery of a novel, potent, and Src family-selective tyrosine kinase inhibitor: study of Lck- and FynT-dependent T cell activation. J. Biol. Chem. 271:695.[Abstract/Free Full Text]
4. Marth, J. D., J. A. Cooper, C. S. King, S. F. Ziegler, D. A. Tinker, R. W. Overell, E. G. Krebs, R. M. Perlmutter. 1988. Neoplastic transformation induced by an activated lymphocyte-specific protein tyrosine kinase (pp56^lck). Mol. Cell. Biol. 8:540.[Abstract/Free Full Text]
5. Cheng, S. H., P. C. Espino, J. Marshall, R. Harvey, J. Merrill, A. E. Smith. 1991. Structural elements that regulate pp59^c-fyn catalytic activity, transforming potential, and ability to associate with polyomavirus middle-T antigen. J. Virol. 65:170.[Abstract/Free Full Text]
6. Veillette, A., M. Fournel. 1990. The CD4 associated tyrosine protein kinase p56^lck is positively regulated through its site of autophosphorylation. Oncogene 5:1455.[Medline]
7. Pingel, J. T., M. L. Thomas. 1989. Evidence that the leukocyte-common antigen is required for antigen-induced T lymphocyte proliferation. Cell 58:1055.[Medline]
8. Koretzky, G. A., J. Picus, M. L. Thomas, A. Weiss. 1990. Tyrosine phosphatase CD45 is essential for coupling T-cell antigen receptor to the phosphatidyl inositol pathway. Nature 346:66.[Medline]
9. Volarevic, S., B. B. Niklinska, C. M. Burns, H. Yamada, C. H. June, F. J. Dumont, J. D. Ashwell. 1992. The CD45 tyrosine phosphatase regulates phosphotyrosine homeostasis and its loss reveals a novel pattern of late T cell receptor-induced Ca²⁺ oscillations. J. Exp. Med. 176:835.[Abstract/Free Full Text]
10. Volarevic, S., B. B. Niklinska, C. M. Burns, C. H. June, A. M. Weissman, J. D. Ashwell. 1993. Regulation of TCR signaling by CD45 lacking transmembrane and extracellular domains. Science 260:541.[Abstract/Free Full Text]
11. Hovis, R. R., J. A. Donovan, M. A. Musci, D. G. Motto, F. D. Goldman, S. E. Ross, G. A. Koretzky. 1993. Rescue of signaling by a chimeric protein containing the cytoplasmic domain of CD45. Science 260:544.[Abstract/Free Full Text]
12. Mustelin, T., K. M. Coggeshall, A. Altman. 1989. Rapid activation of the T-cell tyrosine protein kinase pp56^lck by the CD45 phosphotyrosine phosphatase. Proc. Natl. Acad. Sci. USA 86:6302.[Abstract/Free Full Text]
13. Mustelin, T., T. Pessa-Morikawa, M. Autero, M. Gassmann, L. C. Andersson, C. G. Gahmberg, P. Burn. 1992. Regulation of the p59^fyn protein tyrosine kinase by the CD45 phosphotyrosine phosphatase. Eur. J. Immunol. 22:1173.[Medline]
14. Cahir McFarland, E. D., T. R. Hurley, J. T. Pingel, B. M. Sefton, A. Shaw, M. L. Thomas. 1993. Correlation between Src family member regulation by the protein-tyrosine-phosphatase CD45 and transmembrane signaling through the T-cell receptor. Proc. Natl. Acad. Sci. USA 90:1402.[Abstract/Free Full Text]
15. Sieh, M., J. B. Bolen, A. Weiss. 1993. CD45 specifically modulates binding of Lck to a phosphopeptide encompassing the negative regulatory tyrosine of Lck. EMBO J. 12:315.[Medline]
16. Ostergaard, H. L., D. A. Shackelford, T. R. Hurley, P. Johnson, R. Hyman, B. M. Sefton, I. S. Trowbridge. 1989. Expression of CD45 alters phosphorylation of the lck-encoded tyrosine protein kinase in murine lymphoma T-cell lines. Proc. Natl. Acad. Sci. USA 86:8959.[Abstract/Free Full Text]
17. Hurley, T. R., R. Hyman, B. M. Sefton. 1993. Differential effects of expression of the CD45 tyrosine protein phosphatase on the tyrosine phosphorylation of the lck, fyn, and c-src tyrosine protein kinases. Mol. Cell. Biol. 13:1651.[Abstract/Free Full Text]
18. Burns, C. M., K. Sakaguchi, E. Appella, J. D. Ashwell. 1994. CD45 regulation of tyrosine phosphorylation and enzyme activity of src family kinases. J. Biol. Chem. 269:13594.[Abstract/Free Full Text]
19. D’Oro, U., K. Sakaguchi, E. Appella, J. D. Ashwell. 1996. Mutational analysis of Lck in CD45-negative T cells: dominant role of tyrosine 394 phosphorylation in kinase activity. Mol. Cell. Biol. 16:4996.[Abstract]
20. Chan, A. C., D. M. Desai, A. Weiss. 1994. The role of protein tyrosine kinases and protein tyrosine phosphatases in T cell antigen receptor signal transduction. Annu. Rev. Immunol. 12:555.[Medline]
21. Byth, K. F., L. A. Conroy, S. Howlett, A. J. Smith, J. May, D. R. Alexander, N. Holmes. 1996. CD45-null transgenic mice reveal a positive regulatory role for CD45 in early thymocyte development, in the selection of CD4⁺CD8⁺ thymocytes, and B cell maturation. J. Exp. Med. 183:1707.[Abstract/Free Full Text]
22. Cheng, H. C., H. Nishio, O. Hatase, S. Ralph, J. H. Wang. 1992. A synthetic peptide derived from p34^cdc2 is a specific and efficient substrate of src-family tyrosine kinases. J. Biol. Chem. 267:9248.[Abstract/Free Full Text]
23. Mustelin, T., A. Altman. 1990. Dephosphorylation and activation of the T cell tyrosine kinase pp56^lck by the leukocyte common antigen (CD45). Oncogene 5:809.[Medline]
24. Kmiecik, T. E., D. Shalloway. 1987. Activation and suppression of pp60^c-src transforming ability by mutation of its primary sites of tyrosine phosphorylation. Cell 49:65.[Medline]
25. Weiss, A.. 1998. T lymphocyte activation. W. E. Paul, ed. Fundamental Immunology 4th Ed.411. Lippincott-Raven, Philadelphia.
26. Siegel, J. N., C. H. June. 1996. Signal transduction and T lymphocyte activation. R. R. Rich, and T. A. Fleisher, and B. D. Schwartz, and W. T. Shearer, and W. Strober, eds. Clinical Immunology: Principles and Practice 192. Mosby-Year Book, St. Louis, MO.
27. Biffen, M., D. McMichael-Phillips, T. Larson, A. Venkitaraman, D. Alexander. 1994. The CD45 tyrosine phosphatase regulates specific pools of antigen receptor-associated p59^fyn and CD4-associated p56^lck tyrosine in human T-cells. EMBO J. 13:1920.[Medline]
28. Deans, J. P., S. B. Kanner, R. M. Torres, J. A. Ledbetter. 1992. Interaction of CD4:lck with the T cell receptor/CD3 complex induces early signaling events in the absence of CD45 tyrosine phosphatase. Eur. J. Immunol. 22:661.[Medline]
29. Ostergaard, H. L., I. S. Trowbridge. 1990. Coclustering CD45 with CD4 or CD8 alters the phosphorylation and kinase activity of p56^lck. J. Exp. Med. 172:347.[Abstract/Free Full Text]
30. Imbert, V., J. F. Peyron, D. Farahi Far, B. Mari, P. Auberger, B. Rossi. 1994. Induction of tyrosine phosphorylation and T-cell activation by vanadate peroxide, an inhibitor of protein tyrosine phosphatases. Biochem. J. 297:163.
31. Imbert, V., D. Farahifar, P. Auberger, D. Mary, B. Rossi, J. F. Peyron. 1996. Stimulation of the T-cell antigen receptor-CD3 complex signaling pathway by the tyrosine phosphatase inhibitor pervanadate is mediated by inhibition of CD45: evidence for two interconnected Lck/Fyn- or zap-70-dependent signaling pathways. J. Inflamm. 46:65.[Medline]
32. Roach, T., S. Slater, M. Koval, L. White, E. C. McFarland, M. Okumura, M. Thomas, E. Brown. 1997. CD45 regulates Src family member kinase activity associated with macrophage integrin-mediated adhesion. Curr. Biol. 7:408.[Medline]
33. Stone, J. D., L. A. Conroy, K. F. Byth, R. A. Hederer, S. Howlett, Y. Takemoto, N. Holmes, D. R. Alexander. 1997. Aberrant TCR-mediated signaling in CD45-null thymocytes involves dysfunctional regulation of Lck, Fyn, TCR-, and ZAP-70. J. Immunol. 158:5773.[Abstract]

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				C. Zhou, A. Saxon, and K. Zhang Human Activation-Induced Cytidine Deaminase Is Induced by IL-4 and Negatively Regulated by CD45: Implication of CD45 as a Janus Kinase Phosphatase in Antibody Diversification J. Immunol., February 15, 2003; 170(4): 1887 - 1893. [Abstract] [Full Text] [PDF]

				E. L. Virts, O. Diago, and W. C. Raschke A CD45 minigene restores regulated isoform expression and immune function in CD45-deficient mice: therapeutic implications for human CD45-null severe combined immunodeficiency Blood, February 1, 2003; 101(3): 849 - 855. [Abstract] [Full Text] [PDF]

				A. Harashima, M. Suzuki, A. Okochi, M. Yamamoto, Y. Matsuo, R. Motoda, T. Yoshioka, and K. Orita CD45 tyrosine phosphatase inhibits erythroid differentiation of umbilical cord blood CD34+ cells associated with selective inactivation of Lyn Blood, December 15, 2002; 100(13): 4440 - 4445. [Abstract] [Full Text] [PDF]

				B. Barbeau, G. A. Robichaud, J.-F. Fortin, and M. J. Tremblay Negative Regulation of the NFAT1 Factor by CD45: Implication in HIV-1 Long Terminal Repeat Activation J. Immunol., September 1, 2001; 167(5): 2700 - 2713. [Abstract] [Full Text] [PDF]

				N. Blank, M. Kriegel, T. Hieronymus, T. Geiler, S. Winkler, J. R. Kalden, and H.-M. Lorenz CD45 Tyrosine Phosphatase Controls Common {{gamma}}-Chain Cytokine-Mediated STAT and Extracellular Signal-Related Kinase Phosphorylation in Activated Human Lymphoblasts: Inhibition of Proliferation Without Induction of Apoptosis J. Immunol., May 15, 2001; 166(10): 6034 - 6040. [Abstract] [Full Text] [PDF]

				J. E. Baker, R. Majeti, S. G. Tangye, and A. Weiss Protein Tyrosine Phosphatase CD148-Mediated Inhibition of T-Cell Receptor Signal Transduction Is Associated with Reduced LAT and Phospholipase C{gamma}1 Phosphorylation Mol. Cell. Biol., April 1, 2001; 21(7): 2393 - 2403. [Abstract] [Full Text]

				F. Van Laethem, E. Baus, L. A. Smyth, F. Andris, F. Bex, J. Urbain, D. Kioussis, and O. Leo Glucocorticoids Attenuate T Cell Receptor Signaling J. Exp. Med., March 26, 2001; 193(7): 803 - 814. [Abstract] [Full Text] [PDF]

				S. Trop, J. Charron, C. Arguin, and P. Hugo Thymic Selection Generates T Cells Expressing Self-Reactive TCRs in the Absence of CD45 J. Immunol., September 15, 2000; 165(6): 3073 - 3079. [Abstract] [Full Text] [PDF]

				M. A. Basson, T. J. Wilson, G. A. Legname, N. Sarner, P. D. Tomlinson, V. L. J. Tybulewicz, and R. Zamoyska Early Growth Response (Egr)-1 Gene Induction in the Thymus in Response to TCR Ligation During Early Steps in Positive Selection Is Not Required for CD8 Lineage Commitment J. Immunol., September 1, 2000; 165(5): 2444 - 2450. [Abstract] [Full Text] [PDF]

				K. G. Johnson, S. K. Bromley, M. L. Dustin, and M. L. Thomas A supramolecular basis for CD45 tyrosine phosphatase regulation in sustained T cell activation PNAS, August 29, 2000; 97(18): 10138 - 10143. [Abstract] [Full Text] [PDF]

				R. J. Petrie, P. P. M. Schnetkamp, K. D. Patel, M. Awasthi-Kalia, and J. P. Deans Transient Translocation of the B Cell Receptor and Src Homology 2 Domain-Containing Inositol Phosphatase to Lipid Rafts: Evidence Toward a Role in Calcium Regulation J. Immunol., August 1, 2000; 165(3): 1220 - 1227. [Abstract] [Full Text] [PDF]

				Y. Wang, L. Liang, and W. J. Esselman Regulation of the Calcium/NF-AT T Cell Activation Pathway by the D2 Domain of CD45 J. Immunol., March 1, 2000; 164(5): 2557 - 2564. [Abstract] [Full Text] [PDF]

				E. Rollet-Labelle, C. Gilbert, and P. H. Naccache Modulation of Human Neutrophil Responses to CD32 Cross-Linking by Serine/Threonine Phosphatase Inhibitors: Cross-Talk Between Serine/Threonine and Tyrosine Phosphorylation J. Immunol., January 15, 2000; 164(2): 1020 - 1028. [Abstract] [Full Text] [PDF]

				A Petrone and J Sap Emerging issues in receptor protein tyrosine phosphatase function: lifting fog or simply shifting? J. Cell Sci., January 7, 2000; 113(13): 2345 - 2354. [Abstract] [PDF]

				P. W. Janes, S. C. Ley, and A. I. Magee Aggregation of Lipid Rafts Accompanies Signaling Via the T Cell Antigen Receptor J. Cell Biol., October 18, 1999; 147(2): 447 - 461. [Abstract] [Full Text] [PDF]

				J. R. Seavitt, L. S. White, K. M. Murphy, D. Y. Loh, R. M. Perlmutter, and M. L. Thomas Expression of the p56lck Y505F Mutation in CD45-Deficient Mice Rescues Thymocyte Development Mol. Cell. Biol., June 1, 1999; 19(6): 4200 - 4208. [Abstract] [Full Text] [PDF]

E. D. Sheets, D. Holowka, and B