All ETDs from UAB

Advisory Committee Chair

Harry Schroeder

Advisory Committee Members

Peter Burrows

John Kearney

Paul Geopfert

Randall Davis

Document Type

Dissertation

Date of Award

2012

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Developing B cells exhibit sequential changes in the representation of immunoglobulin (Ig) heavy (H) chain sequences with excess hydrophobic or charged amino acids in complementarity determining region 3 (CDR-H3). To begin to elucidate the mechanisms by which this process occurs, we used Bromodeoxy Uridine (BrdU) incorporation and Annexin V binding to quantify B cell turnover and cell cycle progression in developing B cell subsets in the bone marrow, spleen and peritoneal cavity of wild-type BALB/c mice. To magnify the effects of hydrophobicity and charge, we included in this analysis mice limited to the use of an IgH allele with a single DH gene segment biased in incorporation of neutral, charged, or hydrophobic amino acids; as well as mice lacking the lambda 5 component of the surrogate light chain (SLC). We found that B cells expressing Igs with hydrophobic CDR-H3s exhibited reduced turnover and inefficient cell cycle progression in the transition from the early to late pre-B cell stage, a developmental checkpoint that requires the association of the H chain with SLC to create the pre-B cell receptor. The pattern of cell loss at these stages was similar to that observed in cells lacking lambda 5, and thus constitutionally unable to generate a pre-B cell receptor. B cells with charged or hydrophobic CDR-H3s also exhibited increased loss in the transition from late pre-B to immature B; as well as in the transitions from immature B to mature B in the spleen. In the peritoneal cavity B cells with hydrophobic CDR-H3s exhibited normal turnover, whereas those with charged CDR-H3s contents exhibited increased loss. These findings provide new insight into the mechanisms that limit the production of broadly neutralizing antibodies HIV whose CDR-H3s are enriched for patches of charged or hydrophobic amino acids as well as into the mechanisms that limit the production of highly charged CDR-H3s that are commonly found in pathogenic dsDNA binding antibodies in systemic lupus erythematosus.

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