Distorted antibody repertoire developed in the absence of pre-B cell receptor formation

Abstract

The pre-B cell receptor (pre-BCR), consisting of the μ heavy chain (μHC) and the surrogate light chain (SLC, Vpre-B and λ5), plays important roles during B cell development. The formation of the pre-BCR, which enables the nascent immunoglobulin HC to associate with the SLC, is considered a prerequisite for B cell development. However, a significant number of peripheral mature (leaky) B cells exist in SLC-deficient mice. These leaky B cells develop in the absence of pre-BCR and do not undergo the pre-BCR checkpoint. The antibody repertoires of leaky B cells thus reflect the absence of pre-BCR function. To investigate how the absence of the pre-BCR is circumvented by these leaky-B cells and examine the effect of the pre-BCR checkpoint on the antibody system, we analyzed the antibody repertoires of λ5-deficient (λ5) mice using next-generation sequencing. In λ5 mice, spleen B cells displayed different patterns of VDJ-usage, relative to those in wild-type (WT) mice. Moreover, leaky B cells were neither derived from unusual B2 cells, characterized by particular LC gene rearrangements in the absence of pre-BCR signaling, nor from B1 cells, originating from different B cell progenitors. Analysis of the CDR-H3 amino acid sequences of μ-chain repertoires revealed that certain bone marrow B cells with particular CDR-H3 profiles undergo clonal expansion in λ5 mice. Part of these CDR-H3s contain arginine(s) in the middle of the CDR-H3 loop in λ5 mice, whereas few arginine(s) exist in this middle loop in WT CDR-H3s in the absence of clonal expansion. This CDR-H3 feature in λ5 mice presumably reflects the role of the pre-BCR in autoantibody regulation, since arginine(s) are often found in the antigen-binding site of autoantibodies. Here, we present a unique viewpoint on the role of pre-BCR, by assessing the whole antibody repertoire formed in SLC-deficient mice.

The Role of the Pre-B Cell Receptor in B Cell Development, Repertoire Selection, and Tolerance

“…Nevertheless, the autoimmune reactions include spontaneous formation of T-cell dependent germinal centers, memory B cells and plasma cells that secrete autoantibodies, typical of those found in lupus (SLE), e.g., anti-DNA and anti-nuclear antibodies (ANAs) ( 94 , 95 ). Whether this is unique to mice lacking the entire SL chain is unclear, although splenic B cells in λ5T mice are also enriched for those expressing IgH chains with an arginine-rich H-CDR3 ( 98 ), and more recent work has shown that also λ5T mice secrete autoantibodies ( 90 ). In SLC −/− mice a subset of B cells resembles memory B cells expressing low levels of CD21 (CD21 −/low ) ( 99 ).…”

A Germline-Encoded Structural Arginine Trap Underlies the Anti-DNA Reactivity of a Murine V Gene Segment

“…The URs are non-Ig domains of the two noncovalent bonded polypeptides, VpreB and λ5, which constitute the SLC [ 12 , 13 , 14 ]. The SLC is only expressed in a limited temporal stage of B cell development and may be involved in IgH allelic exclusion [ 15 ], downregulation of recombinant mechanisms [ 16 ], and functionally testing the μH chain, consequently shaping IgH repertoire [ 17 ].…”

Identification of Mouse and Human Antibody Repertoires by Next-Generation Sequencing

“…It is highly potent against invading pathogens due to its vast diversity, fine antigen recognition specificity, and the clonal expansion of antigen-specific B cells. The repertoire of antibody-producing B cells is estimated to be more than 10 15 in a single individual 1 . This immense diversity is generated with the help of VDJ gene recombination in the immunoglobulin genetic loci 2 .…”

“…. Interestingly, IGHV11-2 was also the most common major repertoire in our previously published analysis of naive IgM The method described here is useful for deciphering antigen-responsive antibody repertoires by inclusively analyzing the antibody-repertoire space generated in individual bodies, avoiding inadvertent omission of key antibody repertoires 8,10 . This method also allows the examination of detailed antibody network dynamism, which would facilitate accelerated discovery of protective antibodies against newly emerging pathogens.…”

“…The complementarity-determining region 3 (CDR3) sequences of these productive reads, which give rise to the majority of antigen-binding specificity, are given in IgBLAST outputs. The CDR3 sequences can be analyzed statistically, including biological or technical replicates, as described previously 8,10 .…”