Molecular Interactions in B cell Development

Tufts University Boston, Boston MA

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Abstract

[unreadable] DESCRIPTION (provided by applicant): Maintenance of circulating antibodies requires proper B cell development. Gene rearrangement and editing provide for antibody diversity, but can also result in improperly formed antibody chains. There is a proofing mechanism at the pre-B cell stage of development in which a light-chain like molecule called the surrogate light chain probes the nascent heavy chain for proper folding. Successful complex formation results in assembly of the pre-B cell receptor that ensures survival and proliferation of that particular B cell. The mechanism by which the one and only surrogate light chain assembles and is able to pair with heavy chains is not known. The surrogate light chain comprises two polypeptides, VpreB and 14.1. The hypothesis that the two polypeptides of the surrogate light chain assemble to form a structure that resembles, in part, a mature light chain. Specific aim 1 is three-dimensional structure determination of the variable domain of the surrogate light chain. Using NMR methods we will determine the structure and identify the residues of the variable domain that interact with heavy chain. Specific Aim 2 focuses on identification of the role of unique, non-immunoglobulin domains in surrogate light chain assembly. The extent to which they participate in surrogate light chain assembly will be tested by measuring affinities of a series of constructs with and without the presence of the unique regions using isothermal titration calorimetry and surface plasmon resonance. The extent to which the unique, non-immunoglobulin domains of VpreB and 14.1 proteins have three-dimensional structure will also be determined. This project represents the first structural information for any component of the surrogate light chain of the pre-B cell receptor. Coupling structural information with other biophysical techniques that measure interactions is expected to provide insight on the mechanism of assembly. This R21 application is exploratory because the question is asked whether or not the unique regions have defined structure, either alone, or in complex with each other. Determining the structures will provide the necessary background for understanding and refining models put forward for signal transduction from the pre-B cell receptor that allow proper B cell formation and immunological competency. [unreadable] [unreadable]