Intrinsic bias and public rearrangements in the human immunoglobulin Vλ light chain repertoire.

The immunoglobulin lambda (IGL) repertoires from two unrelated human blood samples, three NOD-scid-IL2Rγ(null) mice engrafted with human hematopoietic stem cells and two pairs of monozygotic twin blood samples were determined by Roche 454 sequencing to generate a total of about 700 000 IGL sequences. We applied bioinformatic analysis to examine IGL repertoires wherein, surprisingly, 20% of CDR-L3 peptide sequences were 'public' (shared across individuals); moreover, full-length IGL protein sequences (VJ recombinants) were also present in the public domain. Subtle yet significant differences in CDR-L3 nontemplated nucleotide addition, IGL V-gene family usage, and amino-acid composition distinguished the public CDR-L3 groups from the private groups. These data suggest that public CDR-L3 intervals can arise by intrinsic genetic mechanisms irrespective of different B-cell developmental milieu (human versus humanized mouse). Furthermore, the occurrence of identical public IGL protein sequences indirectly suggest the positive selection (evolutionary, somatic or both) of particular IGL chains independent of the immunoglobulin heavy chain.

Complement activation in factor D-deficient mice.

To assess the contribution of the alternative pathway in complement activation and host defense and its possible role in the regulation of systemic energy balance in vivo, factor D-deficient mice were generated by gene targeting. The mutant mice have no apparent abnormality in development and their body weights are similar to those of factor D-sufficient littermates. Complement activation could not be initiated in the serum of deficient mice by the alternative pathway activators rabbit erythrocytes and zymosan. Surprisingly, injection of cobra venom factor (CVF) caused a profound and reproducible reduction in serum C3 levels, whereas, as expected, there was no C3 reduction in factor B-deficient mice treated similarly. Studies of C3 and factor B activation in vitro by CVF demonstrated that in factor D-deficient serum the alpha chain of C3 was cleaved gradually over a period of 60 min without detectable cleavage of factor B. CVF-dependent C3 cleavage in the deficient serum required the presence of Mg(2+), whereas in normal mouse serum the presence of divalent cations was not required. These results suggest that in mouse proteolytic cleavage of factor B by factor D is not an absolute requirement for the zymogen to active enzyme conformational transition of CVF-bound factor B. Kinetics of opsonization of Streptococcus pneumoniae by C3 fragments was much slower in factor D-deficient serum, suggesting a significant contribution of the alternative pathway to antibacterial host defense early after infection.

Regulation of the antibody repertoire through control of HCDR3 diversity.

In man, as in mouse, diversification of the antibody repertoire appears to follow a strict developmental program whereby antigen specificities are serially acquired during ontogeny. When compared to the adult repertoire, the fetal antibody repertoire is highly enriched for polyreactive specificities of low affinity. Although the mechanisms governing the development of this fetal repertoire differ between human and mouse, the composition and structure of the fetal antibodies produced by both species are quite homologous. Specifically, both species use similar V gene segments and restrict the sequence and structure of the third complementarity determining region (HCDR3) of the antibody heavy chain. The precise role that this restriction of the HCDR3 might play in the development of immunocompetence in the human remains to be elucidated.