The Role of CD40–CD40 Ligand (CD154) Interactions
in Immunoglobulin Light Chain Repertoire
Generation and Somatic Mutation
Nancy L. Monson,* Sandra J. Foster,* Hans-Peter Brezinschek,* Ruth I. Brezinschek,*
Thomas Do¨rner,* and Peter E. Lipsky,†
,1
*Department of Internal Medicine and †Harold C. Simmons Arthritis Research Center, University of Texas
Southwestern Medical Center at Dallas, Dallas, Texas 75235
To determine whether CD40 ligation influences the
molecular and selective mechanisms that govern the
development of the human Ig light chain repertoire,
analysis of the V
and V
repertoires of CD19
؉
B cells
obtained from a patient with X-linked hyper IgM syn-
drome (XHIM) and a nonfunctional CD154 was carried
out. The nonproductive V
and V
repertoires were
largely comparable to that of the normals with respect
to V gene and J segment distribution as well as CDR3
length and V
L
J
L
joint complexity. Comparison of the
nonproductive and productive repertoires indicated
that a limited number of V
L
genes were positively and
negatively selected in the XHIM patient. Although mu-
tations were observed in the XHIM V
L
repertoires, the
frequency of mutations was significantly lower than in
normals. Typical targeting of these mutations into
RGYW/WRCY motifs was significantly reduced and
subsequent selection of RGYW/WRCY mutations,
which is normally observed, was not found. These re-
sults indicate that CD40 ligation is not required for
generation of the light chain repertoire, positive selec-
tion of some Vk rearrangements, negative selection of
specific V
L
genes, and some degree of somatic muta
-
tion. Importantly, however, targeting of mutations to
RGYW/WRCY motifs and subsequent selection of these
mutated motifs does not occur in the absence of CD40
ligation.
© 2001 Academic Press
Key Words: B lymphocytes; immunoglobulin; somatic
hypermutation; selection.
INTRODUCTION
Naive B cells that recognize T-dependent antigens
require a cognate interaction with CD4
ϩ
T cells in
order to become activated and mature into memory
cells (1, 2). Engagement of CD40 on these B cells by
CD40 ligand (CD154) expressed by activated T cells is
a critical interaction that occurs during T–B cell col-
laboration and initiates many steps of B cell activation,
including induction of proliferation, differentiation,
and immunoglobulin (Ig) production (3–8). In the ab-
sence of CD154 expression, there is no formation of
germinal centers in secondary lymphoid organs and no
generation of memory B cells that recognize T-depen-
dent antigens (9–12). These observations were initially
described in patients with the primary immunodefi-
ciency disorder, X-linked hyper IgM syndrome (XHIM),
which is caused by mutations in the CD154 gene that
prevent its expression or the function of the expressed
protein (13–16), and subsequently confirmed in mice
depleted of CD40 and/or CD154 by targeted gene dis-
ruption (17–19). Thus, naive B cells from XHIM pa-
tients or CD40 or CD154 knockout mice are unable to
respond to T-dependent antigens and to generate ger-
minal center reactions or memory B cells.
To determine whether CD40 engagement plays a
role in the generation of the Ig repertoire, peripheral B
cells from a patient with XHIM who does not express
CD154 were analyzed in detail using PCR amplifica-
tion of V
L
rearrangements from gDNA of individual B
cells. The V
H
repertoire of this patient has been previ
-
ously described (20), and it was suggested that there
were disruptions in the normal selective mechanisms
that shape the expressed V
H
repertoire after V
H
DJ
H
recombination. Moreover, a low degree of somatic mu-
tation was observed in the XHIM V
H
rearrangements,
although the nature of these mutations was not ex-
plored in detail. The current analysis sought to deter-
mine whether the lack of CD40–CD154 interactions
also influenced the expressed light chain repertoire of
this XHIM patient by altering selection and/or the
impact of somatic hypermutation. The data indicate
that the molecular mechanisms that generate the V
L
repertoire are largely intact, as evidenced by the dis-
tribution of V
L
and J
L
elements in the nonproductive
1
To whom correspondence and reprint requests should be ad
-
dressed at National Institute of Arthritis and Musculoskeletal and
Skin Diseases, Building 10, Room 9N228, 10 Center Drive MSC
1820, Bethesda, MD 20892-1820. Fax: (301) 402-0012. E-mail:
lipskyp@mail.nih.gov.
Clinical Immunology
Vol. 100, No. 1, July, pp. 71–81, 2001
doi:10.1006/clim.2001.5049, available online at http://www.idealibrary.com on
1521-6616/01 $35.00
Copyright © 2001 by Academic Press
All rights of reproduction in any form reserved.
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