ligation of the cd23, p45 (blast-2, ebvcs) antigen triggers the cell-cycle progression of activated...

Eur. J. Immunol. 1986.16: 1075-1080 Role of CD23,p45 in B cell growth 1075

John Gordon’, Martin Rowe’, Leonie Walker’ and Graeme Guy’

Departments of Immunology’, Cancer Studies’ and Biochemistry’, The Medical School, University of Birmingham, Birmingham

Ligation of the CD23,p45 (BLAST-2,EBVCS) antigen triggers the cell-cycle progression of activated B lymphocytes*

CD23,p45 (BLAST-2,EBVCS) is a 45-kDa lineage-restricted antigen which appears on the surface of human B cells shortly after activation. A monoclonal antibody (MHM6) to CD23,p45, as well as a polyclonal rabbit antibody raised against the purified antigen were found to promote DNA synthesis in purified tonsillar B cells which had been activated with phorbol ester. Interleukin 1, which was not, by itself, stimulatory for either resting or activated B cells, significantly augmented the growth- promoting properties of MHM6. Kinetic studies indicated that while MHM6 exerted its influence in early GI, interleukin 1 acted later in the cycle just prior to the entry of cells into S phase. The findings demonstrate a role for CD23,p45 in triggering the progression of activated B lymphocytes through the GI phase of the cell cycle. The possibility that this antigen serves as a receptor for a B cell stimulatory factor is discussed.

1 Introduction

A number of structures have been identified on the surface of resting B cells which deliver signals for initiating their clonal expansion and differentiation. The best characterized of these is immunoglobulin (Ig) which serves to bind antigen and sub- sequently trigger a biochemical messenger cascade through the hydrolysis of inositol phospholipids [ l , 21. More recently, the pan-B antigen, Bp35, and the CR2 receptor (gp140) have been implicated in the activation of B cells from the resting state [3-71. For all three structures, it has been found that their ligation with antibody, particularly when in soluble form, need not lead to a full growth signal but, rather, “prime” cells so that they now succumb to the growth-promoting influence of T cells and monocytes or factors derived from them [8-lo]. These include various “B cell growth factors” (BCGF), interleukin 2 (IL2) and IL1. The details of their action, particularly on human B cells, remain unclear.

Candidate receptors for the factors which supplement initial priming signals include antigens which are absent on resting B cells but appear following their activation. Of the molecules in this category, only the Tac antigen (IL2 receptor) has been assigned an unequivocal role in transmitting growth-enhancing signals to appropriately activated B cells [ll-131. IL2, however, acts relatively late in the pathway to B cell expansion, exerting its influence on cells which are already participating in an active growth cycle [14].

One of the earliest lineage-restricted antigens to appear following the activation of resting B cells is the CD23,p45 antigen. Thorley-Lawson and Mann have shown that B cells

[I 54621

* This work was supported by grants from the Medical Research Council (UK).

Correspondence: John Gordon, Department of Immunology, Medical School, University of Birmingham, Birmingham B15 ZTJ, GB

Abbreviations: TPA: 12-0-tetradecanoylphorbol 13-acetate Ig: Immunoglobulin BCGF: B cell growth factor IL: Interleukin dThd: Thymidine

infected with the Epstein-Barr virus (EBV) begin to express this antigen (which they refer to as “BLAST-2”) within 24 h [15]. The constitutively high expression of CD23,p45 on EBV- transformed cell lines led early workers to believe that its induction was specific for cells harboring the viral genome, hence its designation, by one group, of “EBVCS” (CS for cell surface) [16]. Recent studies of our own have shown that even minimal activators of B cells (e.g. inactivated EBV, soluble anti-Ig, calcium ionophore and low doses of phorbol ester) will induce the appearance of CD23,p45 on the B cell surface by as early as 3-4 h post-stimulation [17, 181.

The rapid appearance of CD23,p45 on the activation of nor- mal B cells and its deregulated expression in the transformed state raised the possibility that this antigen was somehow involved in growth control. In this report we show that ligation of CD23,p45, with either a monoclonal or polyclonal antibody, will drive B cells which have been activated with phorbol ester through to DNA synthesis. The effects of receptor ligation were found to be enhanced by the presence of monocyte-derived IL 1. These observations are compatible with CD23,p45 acting to focus growth-promoting signals to B lymphocytes which have been triggered to enter the cell cycle. The possibility that the natural ligand for CD23,p45 is a T cell- derived stimulating factor is discussed.

2 Materials and methods

2.1 Purification of MHM6 Ig

The production of the MHM6-secreting mouse hybridoma has been detailed elsewhere [19]. Monoclonal Ig was prepared from ascites fluid preparations by protein A affinity chromatography as follows. Three ml of ascites was mixed with 3 ml of 1 M Tris buffer, pH 8.0, and clarified by centrifugation before being applied to a 6-ml volume protein A-Sepharose- 4B column (Sigma, Poole, Dorset, GB) which had been equili- brated with 20 mM Tris buffer, pH 8.0. The column was washed with 30 ml of phosphate-buffered saline (PBS), pH 7.2, and the Ig was eluted with 0.1 M citrate buffer, pH 5.6. The MHM6-containing protein peak was collected into 1 M Tris buffer, pH 8.0, and dialyzed against PBS.

Q VCH Verlagsgesellschaft mbH, D-6940 Weinheim, 1986 00 14-2980/86/0909-1075$02.50/0

1076 J. Gordon, M. Rowe, L. Walker and G. Guy

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Eur. J. Immunol. 1986.16: 1075-1080

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2.2 Preparation of rabbit polyclonal antibody to CD23,p45

A polyclonal antiserum was raised in a Lop doe rabbit by hyperimmunization with p45 antigen (which had been purified by affinity chromatography using MHM6 antibody) from EBV-transformed lymphoblastoid cells and incorporated into artificial liposomes with lipid A according to the method of Casali et al. [20]. The rabbit was primed with about 20 pg p45 in liposomes by i.m. and i.p. injections, and boosted 6 weeks later in a similar manner. Serum was collected at 2, 3 and 4 weeks after boost. The immune serum had a titer of > 2000 in indirect immunofluorescence on CD23 ,p45-positive lymphoblastoid cells and was monospecific for p45 in immunopre- cipitation tests with radiolabeled membrane proteins of the same cells. An IgG fraction was prepared by affinity column chromatography using protein A-Sepharose-4B in the same way as described for MHM6 except that the Ig was eluted from the column with 0.1 M citrate buffer, pH 3.0.

2.3 Preparation of B lymphocytes

Highly enriched populations of B cells were prepared from tonsils by a triple negative-selection cycle of cells binding 2- aminoethylisothiouronium bromide-modified sheep erythrocytes (freshly prepared) as described in detail elsewhere [19]. Such populations typically contained > 98% cells positive for surface Ig and <0.2% cells forming rosettes with modified sheep erythrocytes (i .e. T cells). The majority of experiments described were performed on cells collected beneath a 60% Percoll (Pharmacia, Uppsala, Sweden) gradient which had been constructed with fetal calf serum (FCS) as the diluent [21]. These high density populations were designated “resting B cells” and contained < 0.2% cells positive for nonspecific esterase (i. e. monocytes). In some experiments, populations of intermediate and low buoyant densities were prepared by collecting those cells banding between 55 and 60% and above 55% Percoll, respectively.

2.4 B cell stimulations

All activations were performed in 96-well flat-bottom microti- ter plates (growth area = 0.32 cm’) placed at 37°C in a humidified C02-rich atmosphere. Cells were cultured at 5 X 105/ml in 200 p1 of complete growth medium (10% FCS, 2 mM L-glutamine, antibiotics and 5 X lo-’ M Z-mercapto- ethanol). Additions of antibodies, the phorbol ester 12-0-tetradecanoylphorbol 13-acetate (TPA) and affinity-purified, monocyte-derived IL 1 (Genzyme, Boston, MA) were made as indicated in Sect. 3. The activity of the IL 1 was confirmed in an independent thymocyte co-stimulation assay initiated with a suboptimal dose of phytohemagglutinin (PHA) [22].

2.5 Measurement of B cell activation

RNA and DNA synthesis were measured by pulsing cultures with 50 pl of [3H]uridine or [3H)thymidine ([3H]dThd), respectively, both at 10 pCi/ml = 370 kBq, usually between 48 and 64 h but also at other times as indicated in Sect. 3.3. In kinetic studies, cultures were pulsed over the interval preceeding and up to the times indicated on the figures. The results are given as means of triplicate determinations of the radioactivity incorporated which never varied by more than 10%. RNA content

of individual cells was assessed following the method of Dar- zynkiewicz et al. [23]. Briefly, 3 x lo6 cells were taken from culture by pooling 6 replicate wells, washed in serum-free PBS, permeablized by the addition of Triton X-100 and stained with acridine orange (Polysciences, Inc., Warrington, PA) at a final dye concentration of 13 p ~ . Cells were kept on ice and assessed within 30 min for red fluorescence emission on a FACS IV flow cytometer (Becton Dickinson, Mountain View, CA) fitted with the appropriate filters. Acridine orange binding to RNA emits maximally at 640 nm. The data are presented as histogram displays constructed from 50 000 cells. The DNA content of cells was assessed by staining with propidium iodide in hypotonic solution as described elsewhere [24].

3 Results

3.1 Influence of monoclonal and polyclonal antibodies to CD23,p45 on B cell stimulation

Initial studies using the ascites form of the MHM6 monoclonal antibody revealed variable augmentation of DNA synthesis in B cells which had been activated with TPA. The response was highly dose dependent becoming inhibitory at high concentrations of the MHM6 ascites fluid (Fig. 1). Other, irrelevant ascites fluids failed to augment DNA synthesis but often gave inhibition similar to MHM6 ascites when introduced into culture at high levels. By preparing an IgG fraction of MHM6, a plateau augmentation of TPA-driven DNA synthesis was established at concentrations of antibody of 20 yglml and above. Some increase in DNA synthesis was seen even with

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concentrations of MHM6 IgG of less than 1 kg/ml. IgG from a rabbit antiserum raised against the purified p45 antigen yielded a plateau response at the same level of DNA synthesis as obtained with the monoclonal antibody showing that the stimulatory effect was not peculiar to the MHM6 preparation but could be obtained with other antibodies to CD23,p45. Neither the MHM6 ascites, the IgG monoclonal, nor the rabbit polyclonal antibody increased the low level of background DNA synthesis occurring in high-density B cells which had not been activated with phorbol ester (Fig. 1). IgG from nonim- munized or irrelevantly immunized mice or rabbits failed to increase DNA synthesis either in resting B cells or in cells which had been exposed to TPA. All the following experiments described were performed with the IgG preparation of MHM6.

3.2 Augmentation of MHM6 effects by IL 1

As the B cells used in this study were essentially free of monocytes and monocyte products are known to be necessary for certain B cell activations [lo], we examined the influence of adding IL 1 to cultures stimulated with TPA in the presence of MHM6 IgG. It is clear from Fig. 2 that IL 1 at 1 unit/ml significantly enhanced the MHM6-augmented, TPA-driven DNA synthesis in resting B cells. By itself, IL 1 failed to increase the response to TPA alone or to influence, with or without MHM6, the background DNA synthesis in unstimulated B cells. Also from Fig. 2, it is evident that MHM6, either alone or in combination with IL1, did not synergize with levels of TPA which were themselves totally nonstimulatory for promoting DNA synthesis in resting B cells. This is in marked contrast to our observations on experiments where calcium ionophore was used with TPA to promote B cell growth. Here, optimal DNA synthesis could be induced with levels of TPA as low as 0.1 ng/ml. At these concentrations, TPA alone failed to prompt purified resting B cells out of Go [17, 241.

The results presented in Table 1 show that IL 1 exerted its co- operative effect optimally at concentrations of 0.5 unitdm1 and

401n-\ 30

1%- . -.

8 4 2 1 0.5 0.25 043 0

I P A i n g l m l ) __ e z z .

Figure 2. Influence of IL 1 on cells exposed to different concentrations of TPA in the presence of MHM6 IgG. As for Fig. 1 but MHM6 IgG present at a fixed concentration of 20 &mI with (0) or without (0) IL 1 at 1 unit/ml. The DNA synthesis occurring in the absence of MHM6 is also shown with (m) or without (0) the inclusion of IL 1.

Table 1. Influence of ILl at varying concentrations on the MHM6 augmentation of the TPA response

IL 1 ['HIdThd incorporation (cpm) (unitdml) Control TPA~)

-MHM6 +MHM6 -MHM6 +MHM6

0 274 306 4632 18605 0.13 220 300 4543 19 980 0.25 289 267 4664 26 390 0.5 192 297 5480 34 677 1 364 302 5552 34 538 2 225 218 4893 32 729

a) TPA used at 4 ng/ml; MHM6 at 20 &ml.

above. Increasing the concentration of IL1 did not augment TPA-initiated DNA synthesis when used in the absence of MHM6.

3.3 Kinetics of MHM6 and IL 1 effects

The data presented above could be explained either by an increase in the rate of entry into S phase or by an increased number of cells synthesizing DNA. This section describes experiments designed to distinguish between these alterna- tives. The results from kinetic studies shown in Fig. 3 demonstrate that the augmentation of DNA synthesis due to MHM6 and IL 1 represents a prolongation of RNA synthesis followed by an increase in the peak level of DNA synthesis rather than an accelerated progression of cells into S phase. Cells which had been activated with TPA showed the same level of RNA synthesis over the first 26 h irrespective of whether MHM6 and IL 1 were present. By 44 h and at later times, an increase in RNA synthesis was seen if MHM6 had been included in the activations. Similarly, the effect of IL 1 on MHM6-augmented RNA synthesis was first noted at 52 h of culture and became

1 -r -Oi

1, , , , , o ~ ~ ; ~ ~

44 52 68 76 26 18 26 44 52 68

Hours into cul ture D2*3 9

Figure 3. Kinetics of RNA and DNA synthesis in stimulated cultures. As for Fig. 1 but cultures were pulsed with (a) [3H]uridine or (b) rH]dThd over different times. The results are expressed as the net hourly rate of incorporated radioactivity during a pulse preceeding and up to the times indicated. The levels of RNA and DNA synthesis in control cultures have been deducted and never exceeded 100 cpmlh: (0) TPA at 2 ng/ml; (0) TPA + IL 1 at 1 unitlml; (a) TPA + MHM6 IgG at 20 ~g/ml: (0) TPA + IL 1 + MHM6.

1078 J. Gordon, M. Rowe, L. Walker and G. Guy Eur. J. Immunol. 1986.16: 1075-1080

I 44h 1 66h

m a RNA content (arbllrary unltrl

0 0 12 24 36 48

m Time of addit ion ( h o u r s ]

Figure 4. Effect of delaying the addition of MHM6 and IL 1 on DNA synthesis. As for Fig. 1 with DNA synthesis being assessed between 48 and 64 h. Here, cultures were initiated with either TPA + IL 1 (0, 0) or TPA + MHM6 (M, 0) at the concentrations used in Fig. 3 and then either MHM6 (O), IL1 (I) or control culture medium (0, U) was added at the times indicated (again at the concentrations given in Fig. 3).

particularly marked at later times. The point where IL 1 com- menced its influence on RNA synthesis essentially coincided with the time where cells were beginning to enter S phase and synthesize DNA (Fig. 3).

To clarify further the kinetics of growth promotion by MHM6 and IL1, we studied the effect of delaying the addition of either of the two agents into the culture while determining DNA synthesis at a fixed time after initiating the stimulations. It was found that the addition of MHM6 could be delayed by as much as 24 h after the initiation of culture without reducing DNA synthesis between 48 and 64 h (Fig. 4). After 24 h, and particularly by 36 h, the absence of MHM6 during the preced- ing periods significantly diminished the level of subsequent DNA synthesis. Thus, MHM6 was required at around 24 h to influence maximally the subsequent entry of TPA-activated B cells into S phase. By contrast, the addition of IL 1 could be delayed further, so that even adding it at the start of the tritium pulse reduced DNA synthesis by some 40% only. The results from this particular experiment are consistent with the conclusions drawn from the data presented in Fig. 3, that is,

Figure 5. Influence of MHM6 on RNA content of stimulated cells. Cells were cultured as for Fig. 1 but at the times indicated were taken for assessment of RNA content. The fluorescent emission between 600 and 650 nm from acridine orange bound to RNA was analyzed for 50000 cells: (...) control cells; (---) TPA+ILl; (-) TPA + IL 1 + MHM6. All agents were used at the concentrations given in Fig. 3.

MHM6 exerts its growth-promoting action on TPA-activated cells before that of IL 1.

Table 2. Influence of MHM6 on entry of cells into GI + M and on cell number

Next, we monitored the influence of MHM6 on the RNA content of individual cells activated with TPA. Increasing RNA content is a major feature of cells which have left Go and are transisting the G1 phase of the cell cycle [23]. The results shown in Fig. 5 are from experiments where the effect of MHM6 was determined on cells which had been cultured for various times with TPA and IL 1 together; essentially the same profiles were obtained in the absence of IL 1. It can be noted that a proportion of the cells exposed to TPA failed com- pletely to enter the cycle (i.e. increase their RNA content). MHM6 (with or without IL1) was unable to compensate for this but, rather, prompted cells which had already entered the cycle to a higher content of RNA within G1 (Fig. 5). In the absence of TPA, neither MHM6 nor JL 1, alone or in combination, influenced either the RNA content, DNA content, forward scatter or 90' scatter from purified B cells (results not detailed). To determine whether the combination of MHM6 and IL 1 could take TPA-activated cells beyond S phase, into G2 + M, the DNA content of cultured cells was determined with propidium iodide staining and FACS analysis. In addition, cell counts were performed to see whether division was occurring. From the data presented in Table 2 , it can be seen that while the MHM6/IL1 combination was able to drive a proportion of activated cells through S and into Gz + M there was no significant increase in cell number on either day 3 , 4 or 6. A small increase in cell viability was noted up to day 4 where MHM6 had been included in the cultures.

3.4 Influence of MHM6 and IL 1 on B cells of different buoyant densities

All the experiments described above were performed on defined resting populations of high buoyant densities. In this

Stimula- Day 3 Day 4 Day 6 tions') a) Cells cultured at 5 X lO-*/ml in the pres-

% Cells in ence of IL1 (0.5 unitshnl) plus TPA S G2+M Total Viable Total Viable Total Viable (4 ng/ml) and MHM6 IgG (25 p g h l ) as

indicated. Control 0.3 0.1 41(3) 21(2) nd') ndc) ndc) ndcJ b) Represents means of quadruplicate TPA 6.9 1.3 42(1) 22(2) 44(4) 16(4) 49(4) S(2) samples with standard deviations indi- TPA+MHM6 18.0 7.6 41(2) 31(3) 49(3) 24(6) 52(5 ) 6(2) cated in brackets.

Number of ceWd x

c) nd = Not determined.


Table 3. Influence of MHM6 and IL 1 on cells of different buoyant densities

Additions cpm (3H]dThd incorporated into cells of buoyant density

High” Interned. LOW

Control 430 6 955 9 326 MHM6b) 457 7488 9 841 IL1 422 7 094 a 725 MHM6+ILl 435 16086 13 530

a) High, >a%; Intermediate, 55-60%; Low, <55% Percoll. b) MHM6 used at 20 pg/ml; IL 1 at 1 unitlml.

section we examine the effects of adding MHM6 and IL 1 to I3 cells of intermediate and low buoyant densities. By contra\t with resting populations, where usually < 5% of cells are positive for CD23,p45, the majority of B cells of intermediate and low buoyant densities stain convincingly for this antigen (unpublished observations). Furthermore, while some cells in the lower density fractions possess a Go RNNDNA content, significant numbers can be found at all stages of the cell cycle. This is in marked contrast to high buoyant density populations where essentially all cells are in Go (see, for example, Fig. 5).

In the absence of TPA, no tonsillar B cell population responded by increased DNA synthesis to either MHM6 or IL 1 alone (Table 3). However, both the intermediate and the low buoyant density populations, which were already display- ing some “spontaneous” DNA synthesis, showed a significant elevation of dThd incorporation when MHM6 and IL 1 were added together. The augmentation was somewhat greater for the cells banding at intermediate densities than for those of the lowest buoyant densities. For comparison, in independent experiments where medium conditioned by T cells stimulated with PHA and TPA was used as a source of “BCGF 11” [25], DNA synthesis in low buoyant density B cells was augmented by > 10-fold whereas in cells of intermediate density the increase was less than 3-fold (data not detailed). Thus, MHM6, even when used in combination with IL1, acts in a manner which is qualitatively quite distinct from that expected of a BCGF 11.

4 Discussion

For resting murine B cells triggered via their receptors for antigen, a minimum of two factors are required to assist their progression through the cell cycle and into S phase. One of these is the T cell-derived stimulatory factor BSF-1 which appears to exert its influence, at least in part, through priming B cells in Go to respond more vigorously to stimulation by anti- Ig or antigen [26]. The second is the monocyte-derived IL1 which has been shown to act relatively late in the cycle, just prior to the entry of cells into S phase [lo]. Recent studies have confirmed a need for monocyte- and T cell-derived factors in maintaining the replicative cycle of murine B cells which are already proliferating. Here, the details appear somewhat different with monocyte factors acting early in GI while T cell products are required in the G2 phase of the cell cycle [27]. Present understanding of the factors influencing human B cell growth and proliferation is less clear. BCGF I, a

low molecular weight BCGF, has been described which has some similarity with murine BSF-1 [28]. Current evidence, however, suggests that BCGF I acts on cells which have already been primed to enter G1 rather than on the resting (Go) B cell [29]. One study indicates that IL 1 provides a mod- est growth support to such an activation of human B cells [30]. Other T cell products are then believed to act on cells which are already cycling. These include IL2 and BCGF I1 [14, 251.

Much of the confusion surrounding human B cell activation reflects both the paucity of homogeneous growth factors and the use of heterogeneous target B cells. The studies described in this report represent an attempt to clarify some of the details through the use of defined resting populations coupled with an investigation of a cell-surface molecule which may be involved in focusing one of the signals necessary for cell-cycle progression, Resting cells were first prompted into cycle by phorbol ester at a relatively high dose. Kinetic studies then showed that ligation of CD23,p45 conveyed a progression signal to the TPA-activated cells which had entered and become arrested in the early G1 phase of the cell cycle. Although some cells continued to enter S phase, the addition of purified IL 1 significantly increased DNA synthesis in MHMQaugmented cultures. The ability of MHM6 alone to trigger limited DNA synthesis in GI-arrested cells raises the possibility that some IL 1 was produced endogeneously in culture. This could reflect the presence of a few contaminating monocytes or, alterna- tively, production of ILl-like activities by the stimulated B cells themselves [31, 321. It was of interest that populations of intermediate buoyant density, which already contain some “pre-activated” cells, showed, in the absence of TPA, increased DNA synthesis in response to MHM6 only when IL 1 was present. Taken together, these observations suggest that the interaction between MHM6 and IL 1 in signalling the S phase entry of activated B cells might be one of complete synergy. Indeed, the kinetic studies were totally consistent with the notion that IL1 acted on cells which had been triggered to progress through G1 by MHM6. Although such cells continued through to the G2 + M stage of cycle, they did not significantly increase in number. It will be of considerable interest to identify the factors which provide the signals for replication in MHM6-driven cultures.

While we still do not know the nature of the physiological ligand for CD23,p45, the functional studies are compatible with a BCGF I-like activity. It is unlikely to be a BCGF I1 as ligation of the receptor affected cells which were in early GI, rather than the already cycling cells which predominate in the low buoyant density fractions and which represent the major targets for BCGF 11 [33]. It is intriguing that the murine Lyb-2 antigen is both a 45-kDa molecule and that its ligation trans- mits a growth signal to resting B cells [34-]. Given the appar- ent discrepancy in the target cells for human BCGF I and those for murine BSF-1, a functional relationship between CD23,p45 in man and Lyb-2 in mouse may exist. We are cur- rently surveying sources of growth factors which might com- pete with MHM6 and other anti-CD23,p45 antibodies for binding to their target antigen. Preliminary findings show that pre-incubation of cells positive for CD23,p45 with medium conditioned by PHA-activated T cells (a source of BCGF I) inhibits subsequent binding of MHM6 by as much as 70% (Gordon, Lam and Ewing, unpublished observations). We are also initiating experiments designed to explore the role of receptor cross-linking in the action of CD23,p45 by preparing Fab’ and F(ab’)z fragments of the MHM6 antibody; it has

1080 J. Gordon, M. Rowe, L. Walker and G. Guy Eur. J. Immunol. 1986.16: 1075-1080

been shown for Lyb-2 that ligation by monovalent antibody is sufficient to trigger the activation of murine B cells [35]. A relationship between CD23,p45 and a 56-kDa antigen defined by the monoclonal antibody AB-1 which inhibits BCGF-dependent B cell stimulations remains to be established [36].

Received February 19, 1986; in revised form April 1, 1986.

5 References

1 Bijsterboch, M. K., Meade, C. J., Turner, G. A. andKlaus, G. G. B., Cell 1985. 41: 999.

2 Guy, G. R., Gordon, J., Michell, R. H. and Brown, G., Biochem. Biophys. Res. Commun. 1985. 131: 484.

3 Clark, E. A , , Shu, G. and Ledbetter, J. A., Proc. Natl. Acad. Sci. USA 1985. 82: 1766.

4 Golay, J. T., Clark, E. A. and Beverley, P. C. L., J . Immunol. 1985. 135: 3795.

5 Tedder, T. F., Boyd, A. W., Freedman, A. S. , Nadler, L. M. and Schlossman, S. F., J . Immunol. 1985. 135: 973.

6 Frade, R., Crevon, M. C., Barel, M., Vazquez, A,, Krikorian, L., Charriaut, C. and Galanaud, P., Eur. J. Immunol. 1985.15: 73.

7 Nemerow, G. F., McNaughton, M. E. and Cooper, N. R., J . Immunol. 1985. 135: 3068.

8 Kehrl, J. H., Muraguchi, A,, Butler, J. L., Falkoff, R. J. M. and Fauci, A. S., Immunol. Rev. 1984. 78: 75.

9 Kishimoto, T., Yoshizaki, K., Kimoto, M., Okada, M., Kuritani, T., Kikutani, H., Shimizu, K., Nakagawa, T., Nakagawa, N., Miki, Y., Kishi, H., Fukunaga, K., Yoshikubo, T. and Taga, T., lmmunol. Rev. 1984. 78: 97.

10 Howard, M. and Paul, W. E., Annu. Rev. Immunol. 1983.1: 307. 11 Ando, I . , Crawford, D. H., Kissonerghis, M. A,, Owen, M. J. and

12 Boyd, A. W., Fisher, D. C., Fox, D. A., Schlossman, S. F. and

13 Lowenthal, J. W., Zubler, R. H., Nabholz, M. and MacDonald,

Beverley, P. C. L., Eur. J . Immunol. 1985. 15: 341.

Nadler, L. M., J. Immunol. 1985. 134: 2387.

H. R., Nature 1985. 315: 669.

14 Mingari, M. C., Gerosa, F., Moretta, A., Zubler, R. H. and

15 Thorley-Lawson, D. A. and Mann, K. P., J . Exp. Med. 1985.162:

16 Kinter, C. and Sugden, B., Nature 1981. 294: 458. 17 Walker, L., Guy, G., Brown, G., Rowe, M., Milner, A. E. and

18 Gordon, J., Walker, L., Guy, G., Brown, G., Rowe, M. and

19 Rowe, M., Hildreth, J. E. K., Rickinson, A. B. and Epstein, M.

20 Casali, P., Sissons, J. G. P., Fujinami, R. S. and Oldstone, M. B.,

21 Gordon, J., Guy, G. and Walker, L., Immunology 1985. 56: 329. 22 Gordon, J., Guy, G. and Walker, L., Immunology 1986. 57: 419. 23 Darzynkiewicz, Z., Sharpless, T., Staiano-Coico, L. and

Melamed, M. R., Proc. Natl. Acad. Sci. USA 1980. 77: 6696. 24 Guy, G. R., Bunce, C. M., Gordon, J., Michell, R. H. andBrown,

G., Scand. J . Imrnunol. 1985. 22: 591. 25 Yoshizaki, K., Nakagawa, T., Fukunaga, K., Kaieda, T., Maru-

yuma, S. , Kishimoto, S. , Yamamura, Y. and Kishimoto, T., J . Immunol. 1983. 130: 1241.

26 Rabin, E. M., OHara, J. and Paul, W. E., Proc. Natl. Acad. Sci. USA 1985. 82: 2935.

27 Melchers, F. and Lernhardt, W., Proc. Natl. Acad. Sci. USA 1985. 82: 7681.

28 Mehta, S . R., Conrad, D., Sandler, R., Morgan, J., Montagna, R. and Majzel, A. L., J . Immunol. 1985.135: 3298.

29 Muraguchi, A., Kehrl, J. H., Butler, J. L. and Fauci, A. S. , J. Immunol. 1984.132: 176.

30 Falkoff, R. J. M., Muraguchi, A,, Hong, J. X., Butler, J. L., Dinarello, C. A. and Fauci, A. S. , J. Immunol. 1983. 131: 801.

31 Matsushima, K., Kuang, Y-D., Tosato, G., Hopkins, S. J. and Oppenheim, J. J . , Cell. Immunol. 1985. 94: 406.

32 Matsushima, K., Procopio, A., Abe, H., Scala, G., Ortaldo, J. R. and Oppenheim, J. J., J . Immunol. 1985. 135: 1132.

33 Kehrl, J. H., Muraguchi, A., Goldsmith, P. K. and Fauci, S. J., Cell. Immunol. 1985. 96: 38.

34 Subbarao, B. and Mosier, D. E., J . Immunol. 1983.130: 2033. 35 Subbarao, B. and Mosier, D. E., J . Exp. Med. 1984. 159: 1796. 36 Jung, L. K. L. and Fu, S . M., J . Exp. Med. 1984.160: 1919.

Moretta, L., Eur. J . lmmunol. 1985. 15: 193.

45.

Gordon, J., Immunology 1986, in press.

Rickinson, A., Immunology 1986, in press.

A., Int. J . Cancer 1982. 29: 373.

J. Gen. Virol. 1981. 54: 161.

ligation of the cd23, p45 (blast-2, ebvcs) antigen triggers the cell-cycle progression of activated...

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