The B-cell antigen receptor complex: structure and signal transduction

B cell special

The B-cell antigen receptor complex: structure and signal transduction Christopher M. Pleiman, Daniele D'Ambrosio and John C. Cambier The specificity of tbe immune response is determined by the interaction between tbe B-cell ,eceptor (BCR) and its cognate structure, antigen. Recent studies have provided considerable insigbt into the compartmentalization of function witbin this extremely versatile betero-oligomeric receptor complex. In tbis article, Cbristopber Pleiman, Daniele D'Ambrosio and ]obn Cambier consolidate new findings regarding BCR structure and signal transduction. truncated DHJHC~ chain on the cell surface (Fig. 1). Stimulation of this molecular complex with anti-ks antibodies leads to an influx of extracellular Ca 2+ with no detectable increase in the generation of inositol (1,4,5)-trisphosphate [Ins(1,4,5)P3]. At the later pre-B-II stage, a functionally rearranged VHDHJHC (~H) chain can be expressed on the surface with ~.5 and VpreB, replacing DHIHC~. Ligation of this molecular complex, which includes Ig'~, leads to mobilization of extra- and intracellular Ca 2+ stores. Ho vever, it is important to note that attempts to find complexes of I~H with surrogate L chain in normal and B-cell-deficient mice have largely been unsuccessfuls. The occurrence and fuaction of Ig-a, both in the p.H complex and in the complex containing DHJHC., is not yet clear. However, it is known that Ig-,~ is expressed at the pre-B-I stage of development, and presumably associates with Ig-B and is functional in signal transduction 9, Further.more, immature B cells fail to develop in mice expressing nonfunctional Ig-oL, which is consistent with its essential participation in signaling progression at the pre-B-cell stage of development ~°. Receptor-mediated induction of Ins(1,4,5)P3 generation is not detectable until cells have developed to the immature B-cell stage, where a functional L chain is expressed. However, the ability of surrogate receptors on the surface of pre-B cells to transduce signals, and the recently demonstrated requirement for surrogate L-chain and w-chain expression during B-cell development n'12, support the contention that this receptor complex may mediate the activation of B-cell maturation. The apparBCR structure and B-cell development B cells originate from pluripotent hematopoietic ent non-antigen specificity of the receptor complex stem cells and differentiate into lg-secreting plasma suggests that the operative ligand in this process is cells through ~ m'.:'!t-stepligand-driven process (reviewed monomorphlc and possibly derived from stromal cellz. in Refs 6,7). One of the earliest differentiative steps in which signal transduction through a BCR-like structure Compartmentalization of signaling function in the appears to be important is at the precursor pre-B-I stage. BCR subunits As noted earlier, the antigen receptor on immature At this stage of B-cell development, D H- and JH-gene fragments have recombined in the Ig H-chain locus, and mature B cells comprises a clonally distributed while L chains remain in germline configuration. membrane-bound sIg that is noilcovale~itly associated Studies using transformed pre-B-cell-like lines indicate with disulfide-linked Ig-oe and lg-13, or Ig-~, heterothat surrogate L-chain molecules k s and V-~eBcan associ- dimers. Ig-~/ is a truncated form of Ig-[3 lacking the ate with each other and form an Ig-like c~mplex with a C-terminus, and is expressed on low density (i.e. not

The response of B cells to pathogens and other foreign substances is mediated, in part, through the cell-surface B-cell receptor (BCR) for antigen. Stimulation of this receptor by paucivalent proteinacious antigens leads to antigen uptake and processing, as well as the priming of cells to present antigen to T cells 1. Signaling through this receptor can also lead to more-profound biological responses, including activation, tolerance and/or differentiation, depending on the nature of the stimulus and the differentiative state of the B cell2-4. Recent studies have shown that the BCR belongs to a class of receptors that includes the T-cell receptor (TCR), as well as receptors for the Fc region of IgE (Fc¢RI) and IgG (FcR'ylIIa) (reviewed in Ref. 5). These receptors are characterized by a complex hetero-oligomeric structure in which ligand binding and signal transduction are compartmentalized into distinct receptor subunits. In the BCR, the !igand-binding portion of the receptor is surface immunoglobulin (sIg), which is a tetrameric complex of Ig heavy (H) chains and light (L) chains. The sIg-associated 'transducer' structure comprises a disulfide-bonded heterodimer(s) of Ig-ct and Ig-[3, which were recently designated CD79a and CD79b, respectively. These proteins are structurally related and are products of the Ig superfamily genes rob-1 and B29, .~nd function in receptor transport and signal transduction. This review will focus on recent progress in the understanding of the BCR structure and signal transduction.

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Fig. 1. Schematic representation of B-cell development and signaling competence, indicating the relative status of the Ig gene rearrangement and the deteaable signal transduction events. Abbreviations: V,, variable; D, diversity; J, Joining; C, constant; H, heavy; L, light; #H, functionally rearranged sIgM heavy chain; Ins(1,4,j)Ps, inositol (1,4,5)-trisphosphate; slg, surface hnmunoglobulin. in GO) splenic B cells and bone marrow cells. Despite some differences in the glycosylation statu:; of Ig-cx chains, all classes of slg associate with appare;~tly identical Ig-a and Ig-~ heterodimers. Although the. ratio of associated heterodimer to sIg is unknown, the requirement of Ig=a and Ig-lBexpressk, n for sIg transport indicates the ratio is at least 1:1. The bilateral symmetry of s!g suggests a ratio o¢ 2:1. Studies using chimeric proteins, in which the cytoplasmic tail of BCR subunits is expressed with 'inert' transmembrane and extracellular domains, have begun to localize transducer functions within the receptor complex. These studies indicate that Ig-~ and Ig-13cytoplasmic tails, but apparently not izH cytoplasmic tails, possess independent signaling function. However, the sIg

H chain does appear to be able to interact with cytoskeletal elements t3. Furthermore, Ig-- and Ig-L8possess distinct signal transducing capabilities wherein !g-cx, but not Ig-[3, is competent to mediate robust protein tyrosine kinase (PTK) activation and interleukin 2 (IL-2) expression (as shown in the B-cell lymphoma A.20) (Refs 14-17). Although the responses appear qualitatively different, both cytoplasmic tails mediate Ca 2+ mobilization responses. Thus, signaling function can be mediated by both chains. The initial clue as to the localization of the sites within Ig-cx and Ig-[3 that mediate receptor interactions with cytoplasmic effectors was provided by Reth TM. It was observed that Ig-cx and Ig-[~ chains contain a sequence motif of approximately 26 amino acids in length within their tails that is also found in the cytoplasmic tails of other transducer chains, including TCR~, TCR~q, CD3e, CD3% CD38, FceRlJ3, Fc¢RI% human Fc3,RIIa and, potentially, CD22 (Ref. 19). This motif has been variably termed the antigen-receptor homology motif 1 (ARH1) (Ref. 20), the antigen recognition activation motif (ARAM) (Ref. 21) and the tyrosine-based activation motif (TAM) (Ref. 22), and is characterized by six conserved amino acids in the sequence D/E-XT-D/E-X2-Y-Xz-L/I-X7-Y-Xz-L/I.The expression of ARH1 motifs fused to inert extracellular and ,ransmembrane domains carries sufficient structural information to initiate signal transduction events, including PTK activation, protein tyrosine phosphorylation, Ca 2+ mobilization, IL-2 production, cytolytic activffy and endocytosis (reviewed in Ref. 23). Phosphorylation of the two tyrosine residues within the ARH1 motif appears to be a critical event in signal transduction through these receptor subunits 24, since even a conservative mutation of these residues (e.g. to phenylalanine) ablates the ability to transduce signals2s-27. However, tyrosine to phenylalanine mutants in IgM-Ig-~ and !~&M-!g-~chimeras have ~en shown to mediate activation of src-family kinases, albeit at a much lower efficiency than wild-~pe chimeras 2s. Recent studies, using cell lines deficient in lg-oq indicate that the deficit in signaling function of single tyrosine to phenylalanine mutants observed with CD8-Ig-~ chimeras can be compensated by pairing with wildtype Ig-[3 (Ref. 29). However, mutation of both tyrosine residues results in the inactivation of the reconstituted BCR, and suggests a dominant role for Ig-~x in signal transduction mediated by the BCR. This finding is consistent with studies in which Ig-oL-deficient mice were reconstituted with the same double tyrosine to phenylalanine Ig-o~ mutants 1°. The development of B cells in these mice is apparently blocked at the pre-Bcell stage, a result that is reminiscent of those obtained with either hs-knockout or i~H-transmembrane-knockout mice lL12, suggesting that Ig-o~ARH1 tyrosine residues play a critical role in mediating B-cell development.

Immunology roaar 3 9 4

Molecular basis of signal transduction The mechanisms by which antigen receptors activate downstream effectors are beginning to yield to study. The earliest detectable biochemical event that follows aggregation of the BCR is an increase in protein tyrosine phosphorylation. This results, at least in part, from the

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B cell special activation of src-family PTKs including p55 blk, p59 ~n, p56 I~k, p53/56 ]yn and, the more-distantly related PTK, p72 ~yk. A number of earlier studies demonstrated that these kinases associate with the BCR (vresumably with nonphosphoryiateJ ~RH1 motifs) in unstimulated cells (reviewed in Ref. 30). In vitro binding studies suggest that the src-family kinases interact with resting receptors primarily via the Ig-a chain 2°, through an association with their first ten N-terminal residues of the kinase3L This interaction occurs in the absence of phosphorylation of the tyrosine residues within the motif32, which may allow the nonligated receptor to carry resting kinase that can be activated quickly upon receptor ligation. These results are consistent with those observed with p59 fr" binding to the cytoplasmic tails of the TCR complex ~, "q, ~/and • chains 33. The differential ability of Ig-a and Ig-13 to bind srcfamily kinases is attributable to a sequence of four amino acids, DCSM, within the ARH1 motif of Ig-a (the equivalently positioned sequencc in Ig-13 is QTAT) (Ref. 32). Switching these sequences exchanges the binding activity of Ig-a and !g-i3. Importantly, studies using switch mutants indicate that DCSM is critical to the transduction of signals by Ig-a that Ic~.d to IL-2 production in A.20 cells (S. Cassard, J.C. Cambier and C. Bonnerot, unpublished). Thus, DCSM seems to determine the ARHl-mediated binding of src-family kinases to the resting receptor, and this function appears to be of paramount importance in receptor signaling. One of the most immediate consequences of BCR ligation is the phosphorylation of Ig-c~ and Ig-[3 ARH1 motifs on tyrosine residues in the amino acid sequences YEGLN and YEDI. Recently, it was shown that phosphorylation of these residues leads to increased binding of src-family kinase (approximately 20-fold), and this triggers kinase activation 32. The enhanced association is attributable to the binding of src~fami!y kinase srchomology domain 2 (SH2) regions to the phosphorylated tyrosine residues in the ARH1 motif, which results in the concomitant loss of binding to the ten N-terminal residues3L Thus, antigen-induced tyrosine phosphorylation of the ARH1 motif apparently leads to a spatial reorientation of the receptor-associated src-family kinases, such that binding changes from N-terminalmediated to SH2-domain-mediated. This reorientation provides a mechanism for kinase de-repression and activation 34,

occur only following receptor ligation, suggesting that phosphorylation of the tyrosine residues in the ARHi motif may be essential for binding 16. Recent studies have confirmed this possibility by showing that p72 ~ykbinds strongly to phosphorylated Ig-a and Ig-J3 ARH1 motif peptldes (S. Johnson, H. Yamamura, T. Kuro'-:.aki and J.C. Cambier, unpublished). This is not surprising in view of recent findings that a phosphorylated Y-X-X-I/L sequence is a predicted binding site for the p72 ~rk SH2 domains 36, and that a closely related T-cell kinase, ZAP-70, binds to the tyrosine-phosphorylated ARH1 motifs in the cytoplasmic tails of the TCR complex 37. That p72 ~yk does not bind to unstimulated chimeric Ig molecules might be attributed to the fact that these molecules lack slgM transmembrane and cytoplasmic tail sequences, which may be critical for mediating binding of p72 ~ykto I~H. Thus, p72 ~ykmay be involved in BCR signaling at two levels: the first involving binding to the resting receptor via slg transmembrane/cytoplasmic regions; and the second involving SH2-mediated binding to phosphorylated Ig-a and Ig-[3. Activation of receptor-associated kinases Activation of receptor-associated kinases appears to be maximal 15-60 seconds after receptor ligation, as determined by phosphorylation of multiple intracellular substrates as well as the :nases themselves, and increased activity of the kinases in in vitro kinase assays 3s,3s-43. Studies using kinase inhibitors indicate that all known downstream signaling events, as well as subsequent biological responses, are dependent upon PTK activity 44-4s. Although the mechanism for srcfamily PTK activation by phosphorylated ARr-I1 motifs has been defined, it is unclear how the initial ARH1 phosphorylation is triggered. However, in addition to an initial report by Hutchcroft et al.a°, several lines of evidence implicate p72 syk as the first kinase activated

of chimeric receptors containing p72 syk cytoplasmic domains, but not p56 lCkor ZAP-70, initiates inductive tyrosine phosphorylation and Ca2+ mobilization47; (2) p72 ~ykappears to associate with slg in the absence of Ig-~ and Ig-I~ (Ref. 35); (3) knockout of p72 syk in a B-cell lymphoma results in virtually total ablation of BCR signaling4S; (4) src-family kin~ses bind to tyrosine-phosphorylated p72 ~ykthrough their SH2 domains, an event that would require previous p72 syk activation (C.M. Pleiman, P. Andre and J.C. Cambier, unpublished); and (5) recent studies indicate that p72 ~yk is activated by TCR ligation of the p561¢k-deficient A role for p72'# The importance of p72 syk in signal propagation J.CaM 1.6 cell line, and that this activation occurs in through the BCR has been suggested due to its associ- the absence of detectable src-family or ZAP-70 kinase ation with the 'resting' receptor complex, and its im- activation 49. Recently, it has been shown that slgM molecules mediate phosphorylation following receptor ligation. Receptor association was originally proposed to occur via mutated within a transmembrane polar patch are unable slgM since, under solubilization conditions that stripped to activate signal transduction s°. This is consistem with away Ig-a and Igq3 chains, p72 ~ykremained associated the hypothesis that the first activated effector, possibly with slg (Ref. 35). However, a different conclusion was p72~ k, associates with slg, and that the polar patch is reached when p72 syk association was investigated with likely to be the site that mediates such binding in restchimeric molecules comprising the extracellular domain ing receptors. Point mutations in ~H (Ser584-oAla, of IgM, the transmembrane domain of CD8, and the Tyr587-oPhe, Thr592-oVal, or Lys597-~Iie) did not cytoplasmic tails of either Ig-ot or Ig-13 (Ref. 15). The affect association of slgM with Ig-~ and Ig-J3 heterointeraction of p72 ~yk with these chimeras appears to dimers, but totally inhibited antigen-induced signal

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Immunology Today

of the mouse major histocompatibility complex (MHC) clas~ II I-Aa molecule (Ref. 51). The peripheral polar parch mutants, Ser584-->Alaand Lys597-->Ile,remained responsive to stimulation with monoclonal antibodies

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B cell special Fig. 2. Models for slg BCR-mediated activation of signal transduction events. (a) Ligation of the BCR by thymus-de1)endent (TD) antigens may initiate the activation of a tzH-associated effector, possibly I)72".'k. Auto1)hosphon, lation may lead to binding of the SH2 domain of src-family kinases, which 1)bos1)horylate (P) Ig-~ and Ig-t8 ARH1 motifs. Dow,s'tream, ,:,Idit~onal src-[amily kinases may be recruited, as well as Shc, Grb2 and Sos1, and the I)2 lr~"pathway may be activated. (b) Cro~slinkmg of the BCR by a thymusinde1)endent (TI) antigen, or with anti-sIg antibodies, may ~ctivate B cells via extensive lattice formation,. This may directly actwate src-family kinases b~" trans-1)hos1)horylation without the need for an u1)stream effector such as 1)72s'L The downstream events of Shc, Grb2, Sos1 and I)21'~ recruitment would then follow as for TD antigens. Abbreviations: slg, surface immunoglobulm; BCR, B-cell receptor; SH2, src-bomology domain 2; ARH1, antigen-rece1)tor homology 1 domain; Shc, SH2 domain-containing adaptor protein: Grb2, growth factor rece1)tor-bound protein 2; PLCy, 1)hos1)holi1)ase Cy; MAPK, mitogen-activated protein kinase; GAP, GTPase-activating protein; PI3-K, 1)hos1)hoinositide3-kinase. (mAbs), thereby defining the central aspect of this region (YSTTVT) as being of p~" ~cular importance in antigen-induced signal transdL~aon. Point mutations in this polar region of the transmembrane domain of slgM have also been described by another group s2. Not surprisingly, these mutant receptors appeared to signal normally when stimulated with polyclonal antilgM antibodies. However, the ability of these mutants to respond to stimulation by antigen or by anti-IgM mAb was not investigated. Although data regarding the mechanism of antigeninduced signal transduction are somewhat fiagmentary, they are consistent with the model depicted in Fig. 2a. According to this model, ligation of the BCR by antigen initiates signaling by activation of a IxH-aasociated effector, potentially p72 Syk,and autophospho :y!ation leading to src-family kinase SH2 binding and- derepression. The src-family kinase then phosphorylates Ig-ot and Ig-I3 ARH1 motif tyrosine residues. This results in recruitment, and/or reorientation, and activation of additional src-family kinase molecules. Phosphorylation of these tyrosine residues also functions in the recruitment of additional p72 ~yk(Ref. 15). The SH2 domain-containing adaptor protein Shc (D. D'Ambrosio and J.C. Cambier, unpublished), and presumably additional SH2-containing proteins, are also recruited to the receptor following ARH1 phosphorylation. It appears likely that recruited Shc and p72 syk are activated by phosphorylation mediated by src-family kinases. In the case of Shc, phosphorylation leads to association with Grb2 and Sos1, and activation of the p21 ~a~ pathway s3, a result that is consistent with the ability of p21 ras to co-cap with sIg (Ref. 54). Although activation of p72 ~yk by autophosphorylation probably occurs48, the phosphorylation of p72 syk by src-family kinases is supported by the requisite expression of p56 kk for ZAP-70 phosphory!ation in Jurkat cells ss. Furthermore, recent findings suggest that, under certain condit;cms, p53/56 Iy" may directly phosphorylate and modulate the activity of p72 syk (Ref. 56). Additional insight regarding the sequence of kinases activated upon BCR ligation derives from studies of the effects of the tyrosine phosphatase CD45 on signaling. In CD45- J558LIxm3 cells, receptor ligation leads to phosphorylation of a limited repertoire of substrates and a more-robust phosphorylation of p72 Syk(Ref. 57) and Ig-ot and Ig-13 than observed in C1245+ coanterparts, suggesting that the kinase responsible for initiating this phosphorylation is not regulated by CD45 (L. Pan, W. Bedzyk, M. Reth and J.C. Cambier. unpublished). The p72 ~yk molecule is a likely caudidate because it lacks the negative tyrosine phosphorylation

Immunology Today

site found in src-family kinases, these being substrates for CD45. However, the model proposed above appears to be inconsistent with the finding that receptor chimeras containing Ig-~, Ig-13, CD3~ or TCR/~ (Refs 14-17,28) are effective transducers when crosslinked by extracellular ligands, and is also inconsistent with the ability of IXH transmembrane point mutants to respond to polyclonal anti-Ix antibodies s°,s2. It is possible that the normal mechanism of receptor activation can be subverted by extensive formation of receptor lattices. Specifically, p72 Sykmay normally function immediately upstream (as well as downstream) of the src-family kinases, such that activation of p72 syk following antigen stimulation leads to phosphorylation of tyrosine residues in ARH! motifs, as ,,;'eli as ".he subsequent recruitment and activation of additional src-family kinases, p72 ~ykand other effectors (Fig. 2a). However, a strong crosslinking stimulus may lead m direct activation of receptor-associated src-family kinases by a p72~Yk-independent pathway (Fig. 2b). This could occur by high-density receptor patching or capping, leading to the activation -f src-family kinases by trans-phosphorylation. These two modes of receptor activation IJll,ValuIu~91k-ally o y I.IIylIILI3-HK-I-}~IILI~IIL

and -independent type II antigens, respectively. Clearly, further studies are necessary to resolve this important issue. Activation of downstream events The relative impact of src-family kinases versus p72 Syk kinases on downstream eveo,ts is suggested by results of experiments using a chicken B lymphoma in which homologous recombination was used to eliminate either the src-family kinase lyn or syk genes. RNA blot analysis revealed no detectable expression of the src-family kinase genes fyn, lyn, hck, yes, blk, lck or src, or the p72srk-related protein ZAP-70, in these cells48. In syk-deficient cells, BCR ligation did not induce detectable tyrosine phosphorylation of phospholipase C¥2 (PLC,/2). Consistent with this effect, no Ins(1,4,5)P~ generation or Ca 2+ mobilization were observed. Stimulation of the BCR on the lyn-deficient cells produced a slow low-amplitude Ca >- response, with Ins(1,4,5)P 3 generation being normal. These results were interpreted by the authors as suggesting that p53/56 ly" and p72 ~yk act independently, such that: p53/56 ira functions to regulate Ca -'+ mobilization through a process independent of Ins(1,4,5)P 3 generation; while p72 ~yk mediates Ins(1,4,5)P 3 production. However, when total protein tyrosine phosphorylation was investigated in these cells, both mutants were able

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B cell special to mount only weak ts'rosine phosphorylation responses and these involved partially distinct substrates. Even additively, the responses did not approximate the response of the parental cell line, suggesting that p72 ~:k and p53/56 l:~ act cooperatively. Therefore, these results are also consistent with an initial action of p72 "k upstream of p53156 l:~. The minimal tyrosine phosphot3"iation observed m syk-deficient cells may reflect p53156 I:~ activation by BCR lattice-driven mechanisms. BCR competence in lyn-defident cells may reflect the effect of a~vation of sIg-associated p72 ~:k alone. Indeed, this is reminiscent of results obtained following ligation of a CD16 (Fc~RIll)-p72 ~:'kchimera that induced tyrosine phosphorylation and Ca "+ mobilization47. A role for src-family kinases in the regulation of downstream signaling events is suggested by the finding that BCR activation leads to kinase association with effectors that include PLC~,2, GTPase-activating protein (GAPh mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI 3-kinase) 39.5s. In the case of PLC~2, MAPK and GAP, the observed associatiov is attributable to the N-terminal ten residues of the src-family kinases. However, P1 3-kinase binding was shown to occur through an interaction with the SH3 domain of the src-family kinases p53/56 b" and p 5 9tvn but, interesnngly, not p55 blk" (Ref. 58). The region within PI 3-kinase responsible for binding src-family kinase has been localized to a proline-rich area found in the regulatory p85 subunit of PI 3-kinase 59. Since PLC'i2, MAPK and GAP have been shown to be tyrosine-phosphorylated following BCR ligation, the association with src-family kinase may function in positioning the molecules for subsequent phosphorylation. However, BCR-mcdiated tyrosine phosphorylation of the PI 3-kinase has not been shown, suggesting that the binding of the src-family kinase SH3 domain may regulate PI 3-kinase function directly. Consistent with this hypothesis, it has been found that PI 3-kinase is activated in vitro by p53156 b" and p59bn SH3 domains and, furthermore, that this interaction is involved in BCR-mediated activation of PI 3-kinase 59. These findings suggest that PI 3-kinase is activated by an ailosteric mechanism involving binding of src-family kinase SH3 domains to the p85 subunit of PI 3-kinase.

7 Melchers, E, Haasner, D., Grawunder, U. et al. (1994) Annu. Rev. lmmunol. 12, 209-226

8 Karasuyama, H., Rolink, A., Shinkai, Y., Young, E, Alt, EW. and Melchers, E (1994) Cell 77, 133-143 9 Sakaguchi, N., Kashiwamura, S., Kimoto, M., Thalmann, P. and Melchers, E (1988) EMBO J. 7, 3457-3464 10 Torres, R.M., Flaswinkel, H., Reth, M. and Rajewsky, K. 11994) J. Cell. Biocbem. (Suppl. 18D), V343 11 Kitamura, D., Roes, J., Kuhn, R. and Rajewsky, K. ( 1991 ) Nature 350, 423-426 12 Kitamura, D., Kudo, A., Schaal, S., Muller, W., Melchers, E and Rajewsk); K. (1992) Cell 69, 823-831 13 Williams, G.T., Peaker, C.J.G., Patel, K.J. and Neuberger, M.S. (1994) Proc. Natl Acad. Sci. USA 91,474-478 14 Kim, K-M., Alber, G., Weiser, P. and Reth, M. (1993) Eur. J. Immunol. 23, 911-916 15 Law, D.A., Chan, V., Datta, S.K. and DeFranco, A.L. (1993) Curt. Biol. 3, 645-657 16 Sanchez, M., Misulovin, Z., Burkhardt, A.L. et al. (1993) J. Exp. Med. 178, 1049-1057 17 Choquet, D., Ku, G., Cassard, S. et al. (1994)J. Biol. Chem. 269, 6491-6497 18 Reth, M. (1989) Nature 338,383-384 19 Leprince, C., Draves, K.E., Geahlen, R.L. et al. (1993) Proc. Natl Acad. Sci. USA 90, 3236-3240 20 Clark, M.R., Campbell, K.S., Kazlauskas, A. et al. (1992) Science 258, 123-126 21 Weiss, A. (1993) Cell 73,209-212 22 Klausner, R.D. and Samelson, L.E. (1991) Cell 64, 875-878 23 Weiss,A. and Littman, D.R. (1994) Cell 76, 263-274 24 Qian, D., Griswold-Prenner, I., Rosner, M.R. and Fitch, EW. (1993)]. Biol. Chem. 268, 4488--4493 25 Kolanus, W., Romeo, C. and Seed. B. (~992) EMBOJ. 11, 4861--4868 26 Letourneur, E and Klausner, R.D. (1992) Science 255, 79-84 27 Romeo, C., Amiot, M. and Seed, B. (1992) Cell 6~, 889-897 28 Burkhardt, A.L., Costa, T., Misulovin, Z., Stealy, B., oozE., j.o. anu l'~ussenzwelg,XI.L. (IYYq) MOl. Led. Biol. 14, 1095-1103 29 Flaswinkel, H. and Reth, M. (1994) EMBO J. 13, 83-89 30 Cambier, J.C., Pleiman, C.M. and Clark, M.R. (1994) Annu. Rev. lmmunol. 12, 457.486 31 Pleiman, C.M., Abrams, C., Timson-Gauen, L.K etal. (1994) Proc. Natl Acad. Sci. USA 91, 4268.4272 32 Clark, M.R., Johnson, S.A. and Cambier, J.C. (1994) EMBO]. 13, lV11-1914 Christopher M. Pleiman, Daniele D'Ambrosio and John 33 Timson-Gauen, L.K., Kong, A-N.T., Samelson, L.E. and Shaw, A.S. (1992) Mol. Cell. Biol. 12, 5438-5446 C. Cambi.er are at the National Jewish Center for 34 Cooper, J.A. and Howell, B. (1993) Cell 73, 1051-1054 lmmunolog3, and Respiratory Medicine, 1400 Jackson 35 Hutchcroft, J.E., Harrison, M.L. and Geahlen, R.L. Street, Dem,e~ CO 80220, USA. (1992) J. Biol. Chem. 267, 8613-8619 36 Songyang, Z., Shoelson, S.E., McGlade, J. et al. (1994) References Mol. Cell. Biol. 14, 2777-2785 1 Cooke. M.P., Heath, A.W., Shokat, K.M etal. (1994) 37 Wange, Ri., Malek, S.N., Desiderio, S. and Samelson, L.E. I. Exp. Med. 179,425-438 (1993) I. Biol. Chem. 268, 19/97-19801 2 Basten. A., Brink, R., Peake, P. et al. ( 1991 ) lmmunol. 38 Burkhardt, A.L., Brunswick, M., Bolen,J.B. and Mond, J.J. Rel: 122, 5-20 (1991) Proc. Natl Acad. Sci. USA 88, 7410-7414 3 Nemazee, D.. Russell, D., Arnold, B. etal. (1991) 39 Yamanashi, Y., Fukui, Y., Wongsasant, W. et al. (1992) Immunol. R el, 122, 117-136 Proc. Natl Acad. Sci. USA 89, 1118-1122 4 Bachmann, M.E, Rohrer, U.H., Kundig, T.M., Burki, K., 40 Hutchcroft, J.E., Harrison, M.L and Geahlen, R.L. Hengarmer, H. and Zinkernagel, R.M. (1993) Science 262, (1991) J. Biol. Chem. 266, 14846-14849 1448-1451 41 Campbell, M.A. and Sefton, B.M. (1990) EMBOJ. 9, 5 Cambier, J.C. and Jensen, W.A. (1993) Curr. Opin. Genet. 2125-2131 De,,. a., 55-63 42 Campbell, M.A. and Sefton, B.M. (1992) Mol. Cell. Biol. 6 Jongstra, J. and Misener, V. (i993) lmmunol. Rev 132, 12, 2315-2321 107-123 43 Gold, M.R., Law, D.A. and DeFranco, A.L. (1990)

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Regulation of B-cell activation by CD45: a question of mechanism Louis B. Justement, Vergil K. Brown and Jiejian Lin Recent studies have demonstrated that the protein tyrosine phosphatase CD45 plays an integral role in regulation of B-cell function. Most notably, expression of this phosphatase is required for activation of B lymphocytes and entry into the cell cycle. Here, Louis Justement and colleagues review current information concerning the function of CD45 in the B cell. The discussion focuses on two questions that are of central importance: what are the physiological substrates for CD45 and how does reversible tyrosine phosphorylation affect their function? The concept that lymphocyte function is controlled by following ligation of the antigen receptor (AgR). The reversible phosphorylation of tyrosine residues is now involvement of CD45 in regulation of T-cell function well established 1. Both in T and B lymphocytes, specific has been covered in recent reviews and will not be disprocesses associated with development, activation and cussed here 4's. Although studies have ccmvincingly differentiation in response to extracellular ligands in- demonstrated that CD45 plays a critical role in B-cell volve the activation of one or more protein tyrosine activation, relatively little is known concerning the kinases (PTKs). In turn, the signals delivered as a result molecular mechanism by which it exerts its effect on of PTK activation lead to a cascade of events that the cell. Several proteins have been identified as potenmediate the appropriate cellular response through tial substrates for CD45 in B cells, and it is important amplification of the original stimulus and the coor- to clarify whether these or others are physiologically dinated regulation of selected signal transduction relevant. The identification of specific substrates for processes. In order for a PTK to elicit the full range of CD45, and delineation of the effect that reversible physiological responses in any given situation, it is logi- phosphorylation has on their function, will greatly cal to propose that it must act in concert with one or facilitate our understanding of how this PTP regulates more protein tyrosine phosphatases (PTPs) (Refs 2,3). the biology of the B cell. An equally important quesThe coordinated actions of PTKs and PTPs ensure that tion, and one that will not be addressed in depth here, the lymphocyte is responsive to the effects elicited by concerns the mechanism by which the enzymatic activity of CD45 is controlled 4. Due to the abundant reversible protein tyrosine phosphorylation. The receptor-like glycoprotein CD45 is expressed in expression of CD45, and its high specific activity, one abundance on T and B cells and constitutes the pre- would predict that the function of this PTP is regulated dominant PTP in the plasma membrane 4's. Recent such that a dynamic equilibrium between phosphorylstudies have implicated CD45 as an important regu- ation and dephosphorylation of substrates can be mainlatory protein that controls activation of T and B cells tained within the cell. Several mechanisms have been © 1994, Elsevier Science Lid 0167-5699/94/$07.00

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