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Department of Immunology, Duke University Medical Center, Durham, NC 27710
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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CD19 is a B lymphocyte-specific member of the Ig superfamily expressed
by early pre-B cells from the time of heavy chain rearrangement until
plasma cell differentiation (5, 6). The 
240-amino acid cytoplasmic
region of CD19 contains nine conserved tyrosine residues (7), some of
which become rapidly phosphorylated following BCR ligation to generate
functionally active SH2-recognition domains that mediate the
recruitment of regulatory molecules to the cell surface. Src-family
PTKs, Fyn, Lyn, and Lck, are present in immunoprecipitated CD19
complexes following BCR ligation (8, 9, 10, 11, 12), which suggests a proximal
role for CD19 in BCR-mediated signaling. CD19 also interacts with
effector molecules downstream of BCR signaling, such as
phosphatidylinositol-3 kinase and the adapter proteins Vav, Cbl, and
Shc (9, 12, 13, 14, 15, 16). Vav tyrosine phosphorylation is significantly
decreased in CD19-deficient B cells following BCR ligation (13).
Collectively, these observations suggest that CD19 is a central
regulatory component upon which multiple signaling pathways converge.
CD19 functions as a costimulatory molecule for the augmentation of B cell proliferation in vitro (3, 4); although, recent studies in mice that lack or overexpress CD19 indicated that CD19 has functions in addition to its costimulatory role (17, 18, 19). B cells from CD19-deficient mice mature normally within the periphery, but are hyporesponsive to most transmembrane signals, including BCR ligation, LPS, and CD40 ligation + IL-4, which leads to significant deficiencies in proliferation, clonal expansion, and differentiation (5, 6, 17, 20, 21). By contrast, B cells from transgenic mice that overexpress CD19 by 3-fold also mature normally within the periphery but are hyperresponsive to transmembrane signals, proliferate at elevated levels, and generate elevated humoral immune responses (5, 6, 17, 18, 20, 21). Their hyperactivity presumably leads to enhanced negative selection in the bone marrow, which results in diminished numbers of B cells in the peripheral pool. In addition, peripheral tolerance is disrupted in mice that overexpress CD19, which results in autoantibody production (6, 22). These and other observations demonstrate that CD19 also serves as a general response regulator for B cells independent of BCR ligation, and that it defines signaling thresholds critical for expansion of the peripheral B cell pool (23).
To further elucidate the molecular mechanisms by which CD19 regulates intracellular signaling pathways in vivo, the activation of signaling molecules following BCR ligation was assessed using primary B cells from CD19-deficient mice. We show a novel functional activity for CD19 wherein it amplifies the kinase activity of Lyn and other Src-family PTKs. The existence of this novel Src-family kinase amplification loop may explain how CD19 regulates basal signaling thresholds and how it augments BCR signaling. Moreover, the absence of this CD19/Src-family kinase amplification loop may further explain why CD19-deficient B cells are hyporesponsive to transmembrane signals.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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All experiments used 2-mo-old CD19-deficient mice (129 x C57BL/6) housed in a specific pathogen-free barrier facility, as described (17). Wild-type littermates generated from heterozygous matings were used as controls. All procedures were approved by the Animal Care and Use Committee of Duke University (Durham, NC). A20 cells were cultured in RPMI 1640 medium (Life Technologies, Gaithersburg, MD) supplemented with 10% FCS, L-glutamine, streptomycin, penicillin, and 2-ME.
Reagents and immunofluorescence analysis
Abs used in this study included: anti-Lyn, anti-Fyn, anti-Blk, anti-Syk, anti-Vav, and anti-Fgr antisera (Santa Cruz Biotechnology, Santa Cruz, CA); monoclonal anti-mouse CD19 Ab (MB19-1) (6); rabbit anti-CD19 cytoplasmic domain antiserum (no. 5382; generously provided by Dr. M. Grove, Duke University); anti-CD79a (MB-1) Ab (generously provided by Dr. L. Matsuuchi, University of British Columbia, Vancouver, Canada); anti-B220 (CD45RA, RA3-6B2; generously provided by Dr. R. Coffman, DNAX, Palo Alto, CA); biotinylated goat anti-mouse IgM Ab (Southern Biotechnology Associates, Birmingham, AL); and F(ab')2 fragments of goat anti-mouse IgM Abs (Cappel, Durham, NC). PE-conjugated streptavidin (Fischer Scientific, Fair Lawn, NJ) was used to reveal biotin-coupled Ab staining.
Immunofluorescence analysis was performed as described (18). Briefly, single cell suspensions were isolated from mouse spleens and counted with a hemocytometer. Leukocytes (106) were stained at 4°C using predetermined optimal concentrations of Abs for 20 min. Cells were washed and analyzed on a FACScan flow cytometer (Becton Dickinson, San Jose, CA).
A cDNA-encoding GST-CD19 fusion protein containing amino acid residues
315–543 (exons 7–14) of mouse CD19 intracellular domain (7) was
expressed in Escherichia coli DH5
and was purified with
glutathione-agarose beads (Pharmacia Biotech, Piscataway, NJ).
SDS-PAGE analysis of purified CD19-GST fusion protein revealed a single
major band of 65 kDa.
B cell activation, immunoprecipitations, and Western blot analysis
B cells were purified from single cell splenocyte suspensions by removing T cells with anti-Thy1.2 Ab-coated magnetic beads (Dynal, Lake Success, NY). B cell suspensions were always >95% B220+, as determined by flow cytometry analysis. B cells were resuspended (2 x 107/ml) into RPMI 1640 medium containing 5% FCS at 37°C. Cells were stimulated with goat anti-mouse IgM Ab F(ab')2 fragments (40 µg/ml; Cappel), and subsequently lysed in buffer containing 1% Nonidet P-40, 150 mM NaCl, 50 mM Tris-HCl (pH 8.0), 1 mM Na orthovanadate, 2 mM EDTA, 50 mM NaF, and protease inhibitors, as described (24). Protein concentrations were determined by light absorbance at 280 nm. For immunoprecipitation, the cell lysates were precleared twice by incubating with appropriate control Abs plus protein A- or protein G-Sepharose beads (Pharmacia), followed by incubating with protein A- or protein G-beads plus rabbit antiserum overnight at 4°C. For CD19 immunoprecipitations, the lysates were precleared with Affigel 10 beads (Bio-Rad, Richmond, CA), conjugated with mouse IgA Ab, then incubated for 3 h with Affigel 10 beads bearing MB19-1 Ab. After washing with lysis buffer four times, immunoprecipitated proteins were subjected to SDS-PAGE with subsequent electrophoretic transfer to nitrocellulose membranes. These membranes were incubated with HRP-conjugated antiphosphotyrosine Ab (4G10; Upstate Biotechnology, Lake Placid, NY) to detect protein phosphorylation, or were incubated with specific Abs against proteins of interest, followed by incubation with HRP-conjugated donkey anti-rabbit IgG Abs (Jackson ImmunoResearch Laboratories, West Grove, PA). These blots were developed using an enhanced chemiluminescence kit (Pierce, Rockford, IL). To verify equivalent amounts of protein in each lane, the blots were stripped and reprobed with Abs against the proteins of interest.
In vitro PTK assays
Splenic B cell lysates were extensively precleared with protein
A-Sepharose beads before immunoprecipitations using anti-Lyn or
anti-Syk Abs and protein A-Sepharose beads for 3 h at 4°C.
The beads were subsequently washed four times in lysis buffer and twice
in reaction buffer (50 mM Tris-HCl (pH 7.4), 10 mM MgCl2,
0.1 mM Na orthovanadate, 1 mM DTT). The beads were then incubated in 50
µl of reaction buffer containing 10 µCi of
[
-32P]ATP for 5 min at 25°C. The reactions were
terminated by adding 50 µl of Laemmli’s 2x sample buffer and
immersion in a boiling water bath for 3 min. The samples were analyzed
by SDS-PAGE with autoradiography. Incorporated 32P was
quantified using a PhosphorImager (Molecular Dynamics, Sunnyvale, CA).
To estimate Lyn activity in the presence of CD19-GST fusion protein,
purified bovine Lyn (5 U; Upstate Biotechnology) was incubated with 0.3
µg of CD19-GST fusion protein or GST in 20 µl of reaction buffer
containing ATP (10 µM) for 10 min at 30°C. The reactions were
terminated by adding 50 µl of 2x sample buffer, and the proteins
were subjected to SDS-PAGE, transferred to nitrocellulose, followed by
immunoblotting with antiphosphotyrosine Ab. Alternatively, cdc2(6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20)
peptide (10 µg) and 10 µCi of [-32P]ATP were added
to the reaction mixture with additional incubation for 10 min. These
reactions were terminated by adding 25 µl of 40% TCA, and a 25-µl
aliquot of the protein precipitate was spotted onto p81
phosphocellulose paper. The phosphocellulose paper was washed five
times with 0.75% phosphoric acid, and once with acetone. Radioactivity
was quantified by scintillation counting. Alternatively, Vav
immunoprecipitated from unstimulated A20 cell lysates (107
cells) was added to the reactions containing purified Lyn and CD19-GST
fusion protein with incubation for 15 min to assess Vav
phosphorylation. The reactions were terminated, and the proteins were
subjected to SDS-PAGE followed by immunoblotting with
antiphosphotyrosine Ab.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The role of CD19 expression in regulating BCR-induced protein
tyrosine phosphorylation was assessed by stimulating purified splenic B
cells from CD19-deficient and wild-type littermates with optimal
concentrations of anti-IgM Abs. Tyrosine phosphorylation of total
cellular proteins was estimated by antiphosphotyrosine immunoblotting
of whole cell lysates generated from B cells at different time points
following BCR cross-linking. Overall, tyrosine phosphorylation was
markedly decreased in B cells from CD19-deficient mice relative to
wild-type littermates before Ag receptor stimulation (Fig. 1
). Following BCR cross-linking, the
overall kinetics of protein tyrosine phosphorylation was similar
between wild-type and CD19-deficient B cells, and tyrosine
phosphorylation of some proteins was normal in CD19-deficient B cells.
However, tyrosine phosphorylation of numerous molecules was
dramatically decreased in CD19-deficient B cells after BCR ligation.
Most notable was decreased phosphorylation of 150-, 80-, 53- to 56-,
45-, and 30-kDa proteins (Fig 1, arrows). The decrease in tyrosine
phosphorylation observed in CD19-deficient B cells is unlikely to
reflect differences in Ag receptor expression, since B cells from
CD19-deficient and wild-type mice expressed similar levels of cell
surface IgM (10 ± 5% difference; Fig. 2A). In addition,
CD19-deficient B cells expressed wild-type levels of Lyn, Fyn, Syk,
Vav, Btk, and phospholipase C-2 proteins (Fig. 2B and
data not shown). Altered maturation of CD19-deficient B cells is
unlikely since CD19-deficient B cells develop normally, are
phenotypically normal (17), and express wild-type levels of Fgr (Fig. 2B), which is predominantly expressed by mature B cells
(25). Therefore, CD19 loss results in decreased tyrosine
phosphorylation of multiple effector molecules downstream of BCR
ligation.
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Signaling molecules affected by the loss of CD19 were identified
by assessing tyrosine phosphorylation of the Src-family PTKs
immunoprecipitated from B cell lysates. Low level phosphorylation of
Lyn, Fyn, Blk, and Lck was evident in B cell lysates from both
CD19-deficient and wild-type littermates (Fig. 3, A–C, and data not shown).
Lyn (Fig. 3A), Blk (Fig. 3C), and Lck (data not
shown) phosphorylation increased in CD19-deficient B cells following
BCR cross-linking, but at lower levels than in wild-type B cells. BCR
ligation did not induce appreciable Fyn phosphorylation in
CD19-deficient B cells when compared with wild-type B cells (Fig. 3B). The kinetics of Src-family PTK phosphorylation was not
measurably altered in multiple experiments (Lyn, 6 experiments; Fyn,
10; Blk, 3; Lck, 2), just their overall phosphorylation levels.
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A). Kinase assays using
cdc2(6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20)NH2 peptide as an exogenous substrate generated
identical results (data not shown). Therefore, CD19 expression
up-regulates BCR-induced activation of the Src-family PTKs.
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C). CD79a protein was expressed at wild-type levels
in CD19-deficient B cells (Fig. 3C). Tyrosine
phosphorylation of 35- to 40-kDa proteins coimmunoprecipitated with
CD79a was also diminished in CD19-deficient B cells after BCR
cross-linking. These proteins are likely to be CD79b, which forms a
heterodimer with CD79a in the BCR complex. Thus, CD19 positively
regulates BCR-induced tyrosine phosphorylation of CD79a and CD79b. CD19 loss does not affect Syk activation
Low level Syk phosphorylation was detected in B cells purified
from spleens of both CD19-deficient and wild-type littermates (data not
shown). After BCR cross-linking, Syk tyrosine phosphorylation in
CD19-deficient B cells was similar to that in wild-type B cells (Fig. 3D). Syk kinase activity was also similar between wild-type
and CD19-deficient B cells before and after BCR ligation, as determined
using autophosphorylation in vitro kinase assays (Fig. 4B).
Therefore, CD19 loss had no discernible affects on Syk activation.
Lyn and Vav interactions with CD19
Since CD19 serves a primary role in Lyn (Fig. 2) and Vav (12, 13, 16) phosphorylation during BCR signal transduction, their associations
with CD19 were assessed in primary mouse B cells. CD19
immunoprecipitated from wild-type splenic B cells before and after BCR
ligation was transferred to immunoblots and probed with
antiphosphotyrosine Abs. Tyrosine-phosphorylated CD19 migrated as a
broad 93- to 100-kDa band that was most easily detected following BCR
ligation (Fig. 5). Immunoblots using a
more sensitive antiphosphotyrosine Ab (PY99; Santa Cruz Biotechnology)
revealed that CD19 was also tyrosine-phosphorylated in unactivated B
cells and that CD19 remained more heavily phosphorylated after 5 min of
BCR ligation than before ligation (data not shown). However, a heavily
phosphorylated 53- to 56-kDa protein doublet coimmunoprecipitated with
CD19 before and following BCR cross-linking. These proteins are likely
to be Src-family PTKs, since reprobing the same Western blot with
anti-Lyn Abs revealed that Lyn was constitutively complexed with
CD19 (Fig. 5). Reprobing the same Western blot with anti-Vav Abs
revealed that Vav also associated with CD19 in unstimulated cells and
that the sharp band comigrating within the broad CD19 band was Vav
(Fig. 5). The associations between CD19, Lyn, and Vav increased after
BCR cross-linking in parallel with increased CD19 phosphorylation
(Fig. 5). These results demonstrate that, although CD19 is not heavily
tyrosine-phosphorylated in resting or activated primary B cells, it
nonetheless associates specifically with Lyn and Vav.
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To determine whether CD19/Lyn interactions affect Lyn activation,
in vitro kinase assays were performed using purified Lyn in the
presence or absence of CD19. Incubation of Lyn with ATP resulted in
autophosphorylation of the single 56-kDa Lyn protein (Fig. 6A). Incubation of a GST
fusion protein encoding the entire CD19 cytoplasmic domain under
identical conditions did not lead to CD19 phosphorylation.
However, adding CD19-GST fusion protein to the Lyn kinase assay
dramatically up-regulated both CD19 and Lyn phosphorylation (Fig. 6A). Incubation of Lyn with GST protein alone did not affect
Lyn autophosphorylation or result in detectable GST protein
phosphorylation.
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-32P]ATP, with or without
cdc2 peptide, resulted in only background levels of incorporated
radioactivity (Fig. 6B). The addition of Lyn and control GST
protein to the in vitro assays resulted in significant Lyn
autophosphorylation and cdc2 peptide phosphorylation (Fig. 6B). However, Lyn kinase activity was dramatically increased
by the addition of CD19-GST fusion protein, which greatly enhanced cdc2
peptide phosphorylation (740% increase, p < 0.05).
Identical experiments using purified Fyn kinase generated essentially
the same results as those shown for Lyn (data not shown). Therefore,
CD19 serves as a substrate for Src-family PTKs, but also amplifies
Src-family PTK activity.
Whether Lyn could phosphorylate Vav, and whether CD19/Lyn interactions
influenced Vav phosphorylation was assessed using the same in vitro
kinase assays. When Vav immunoprecipitated from A20 cells was added to
in vitro kinase assays with CD19-GST fusion protein, neither protein
was phosphorylated (Fig. 6C). The mixture of Vav, Lyn, and
GST protein resulted in phosphorylation of both Vav and Lyn. However,
the addition of CD19-GST fusion protein resulted in significantly
enhanced Vav and Lyn phosphorylation. Thus, Lyn can phosphorylate Vav
directly. Moreover, Lyn phosphorylation of CD19 and the formation of
Lyn/CD19 complexes are likely to enhance both the binding of Vav to
CD19 and Lyn phosphorylation of Vav.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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).
Specifically, tyrosine phosphorylation of Src-family PTKs, Lyn, Fyn,
Blk, and Lck, and CD79a/b was decreased to varying extents following
BCR ligation in the absence of CD19 (Fig. 3, and data not shown). Lyn
kinase activity following BCR cross-linking was also significantly
reduced (Fig. 4A). CD19-deficient B cells also displayed
diminished tyrosine phosphorylation of multiple other proteins that lie
downstream of the Src-family PTKs during BCR signaling, including Vav,
Cbl, and Shc (data not shown, and 13). The inadequacy of
Src-family PTK activation in CD19-deficient B cells correlates with the
observation that CD19 amplified the activity of Lyn and Fyn during in
vitro kinase assays (Fig. 6, and data not shown). That CD19 amplifies
Src-family PTK activation sheds new light on the mechanisms by which
CD19 augments BCR-mediated signaling in murine primary B cells and may
explain the hyporesponsive phenotype of B cells from CD19-deficient
mice.
While Src-family PTK phosphorylation was down-regulated in B cells from
CD19-deficient mice, Syk phosphorylation and kinase activity was normal
(Fig. 3 and 4). Similarly, tyrosine phosphorylation of phospholipase
C-2, which is downstream of Syk, was similar between CD19-deficient
and wild-type B cells (data not shown). Thus, even in the absence of
CD19, BCR ligation leads to low-level activation of Lyn and other
Src-family PTKs, which in turn phosphorylate CD79a and CD79b at levels
adequate for the recruitment of Syk, which binds through its SH2
domains and subsequently becomes phosphorylated (1). Since Src-family
PTKs initiate an activation loop that results in Syk
autophosphorylation (26), wild-type levels of Src-family PTK activation
may not be necessary for optimal Syk activation. Consistent with this,
Syk can be activated in the absence of Lyn (27). Nonetheless, activated
Src-family PTKs or Syk (28) may phosphorylate CD19 on appropriate
residues, which initiates additional rounds of Src-family PTK
activation and amplification of BCR and other signaling cascades
through CD19-dependent processes. Therefore, a critical function for
CD19 appears to be the amplification of Src-family PTK-dependent
signaling cascades, rather than Syk-dependent pathways.
The finding that Lyn’s kinase activity was significantly amplified by
the presence of CD19 during in vitro assays (Fig. 6) provides a direct
mechanism for CD19 function in addition to its previously hypothesized
role as a specialized adapter protein for recruiting signaling effector
molecules (3, 4). CD19 amplification of Src-family PTK function also
provides additional insight into why CD19 supplies potent costimulatory
function when coligated with the BCR complex. However, the finding that
CD19 was associated with both Lyn and Vav in splenic B cells before BCR
ligation suggests that CD19/Lyn/Vav complexes are constitutively
assembled (Fig. 5). Similar to the CD19/Lyn/Vav complex, the BCR is
constitutively organized into a preformed transducer complex in the
absence of Ag ligation (29), as is the T cell Ag receptor complex (30).
The absence of a constitutive CD19/Lyn/Vav signaling complex
potentially explains why B cells from CD19-deficient mice are
hyporesponsive to transmembrane signals. Similarly, the formation of
CD19/Lyn/Vav complexes may be augmented in transgenic mice that
overexpress CD19, which may explain why B cells from these mice are
hyperresponsive to transmembrane signals and are phenotypically similar
to chronically stimulated B cells (5, 6). Constitutive CD19/Lyn/Vav
complex signaling may therefore be responsible for the establishment of
baseline signaling thresholds in B cells before Ag receptor ligation,
in addition to accelerating signaling following BCR engagement or other
transmembrane signals.
In addition to amplifying Src-family PTK activity, multiple findings
suggest that CD19 also facilitates molecular interactions that lead to
Vav phosphorylation (12, 13, 16). Vav and Lyn may share common
signaling pathways regulated by CD19, since tyrosine phosphorylation of
both proteins is decreased in CD19-deficient B cells (Fig. 3, and 13) and Lyn phosphorylated Vav during in vitro kinase assays (Fig. 6C). The abolition of Vav tyrosine phosphorylation in
Lyn-deficient mice also supports this notion (31), in addition to the
findings that Lck and Fyn phosphorylate Vav (32, 33). CD19 may
therefore function as a specialized adapter protein for orchestrating
Lyn/Vav interactions and Vav phosphorylation. That a specific region of
CD19 distinct from the predominant Vav binding region accounts for all
of CD19’s ability to amplify Lyn kinase activity (M. Fujimoto and T.
F. Tedder, manuscript in preparation) suggests that phosphorylated CD19
provides distinct and specific SH2-domain recognition regions to which
Lyn (and other Src-family PTKs) and Vav bind. Src-family PTK binding to
CD19 amplifies PTK activity, which facilitates efficient Vav
recruitment and phosphorylation by the activated PTK. Vav may then
attract other SH2 domain-containing signaling molecules to the CD19
complex, which leads to downstream activation of mitogen-activated
protein kinase cascades (34, 35, 36). The regulation of Src-family PTK
activation and Vav phosphorylation by CD19 thereby provides a potent
molecular mechanism for amplifying BCR signals.
These studies demonstrate a positive regulatory role for CD19 and Lyn in amplifying BCR signal transduction. However, Lyn has both positive and negative roles in signal transduction and participates both in the initiation and termination of BCR-mediated signaling in B lymphocytes (2). Based on the autoimmune phenotype of Lyn-deficient mice, signal termination is a critical function for Lyn (31, 37, 38). Nonetheless, an important consideration is that CD19 expression is B cell-restricted, while Lyn is expressed broadly within the hematopoietic system. In fact, it is likely that dysregulation of Lyn function in non-B lineage cells and resultant systemic inflammation are major causes of autoimmunity in Lyn-deficient mice. Therefore, it is more appropriate to consider that Lyn activity influences an array of signaling pathways, which can have either positive or negative effects. Similarly, CD19 has both positive and negative functions during B cell activation (13, 39). That CD19-deficient mice and Lyn-deficient mice exhibit opposing phenotypes (17, 19, 31, 37, 38) may also be explained by the finding that CD19 expression also regulates the activation of other Src-family PTK members. In addition, Src-family PTK activation was impaired but not completely lost in CD19-deficient B cells. Thus, the collective phenotype of Lyn-deficient mice is likely to reflect multiple qualitative and quantitative factors that are not manifested in CD19-deficient mice.
In summary, these studies demonstrate that CD19 is likely to amplify Src-family PTK activity downstream of BCR ligation. The constitutive assembly of CD19/Lyn/Vav complexes is also likely to influence baseline signaling thresholds. Although Lyn is the most dominant Src-family PTK in B cells (2), CD19 also influenced the activity of other Src-family PTKs. Since CD19 and Lyn have both positive and negative regulatory roles during B cell activation (2, 23, 40, 41, 42, 43), synergistic interactions between these molecules are likely to have complex ramifications. However, the phenotype of CD19-deficient mice is likely to reflect the impaired activity of not only Lyn, but also other Src-family PTKs and their downstream signaling molecules.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Thomas F. Tedder, Department of Immunology, Box 3010, Duke University Medical Center, Durham, NC 27710. E-mail address:
3 Abbreviations used in this paper: BCR, B cell Ag receptor; SH2, Src homology 2; PTK, protein tyrosine kinase.
Received for publication January 22, 1999. Accepted for publication April 5, 1999.
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in murine thymocytes and lymph node T cells. Immunity 1:675.[Medline]
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M. Hasegawa, M. Fujimoto, J. C. Poe, D. A. Steeber, and T. F. Tedder CD19 Can Regulate B Lymphocyte Signal Transduction Independent of Complement Activation J. Immunol., September 15, 2001; 167(6): 3190 - 3200. [Abstract] [Full Text] [PDF]
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