Human B Cell Differentiation in a Model System

Project Details

Description

DESCRIPTION (provided by applicant): We have identified a B-cell immunodeficiency that is distinguished by defective responses to CD4O and IL-4 signaling. B-cells from a young female patient (pt#l) have normal expression of CD4O but are clearly deficient in a subset of CD4O-mediated functions including early signals required for switch recombination. However, under specific in vitro conditions pt#1 B-cells can regain functional responsiveness and undergo switching to express downstream antibody classes or isotypes. Our preliminary data support a model whereby a signaling molecule or transcription factor in the CD4O signal transduction pathway leading to NF-kB activation is affected. This hypothesis is supported by our finding that pt#1 B-cells that are transformed by Epstein-Barr virus (EBV) do not express CD23 a cell surface molecule that is critically dependent on the viral latent infection membrane protein LMP)1 and the subsequent activation of NF-kB by this protein. Furthermore, LMP1 usurps the CD4O signaling pathway in order to maintain cell transformation. Surprisingly, the pt#1 EBV-transformed B-cells loose their transforming potential when grown in dilute culture conditions which also suggests that LMP1 activity is compromised. Thus, three related lines of data strongly suggest that the pt#1 defect is located in the CD4O signaling pathway that leads to NF-kB activation and the transcription of specific cellular genes involved in B-cell activation. We propose to use primary and transformed B-cells from pt#1 to characterize the underlying defect Leading to B-cell dysfunction. Efforts will be initially focused on the characterization of TRAF and NF-kB expression and function which are the most proximal and distal signaling events in the CD4O signaling cascade, respectively. The EBV-transformed pt#l B-cells will be used to analyze signaling of LMP1 via the CD4O pathway. Also, these cells will be used in transfection studies that are aimed at complimenting the pt#l defect and in the analysis of of NF-kappaB responsive promoters. Characterization of signaling pathways under the different growth conditions will provide information into the relationship between these signaling pathways and cell transformation. Finally, pt#l T-cells have subtle defects in helper function provided to control B-cells that manifest in inappropriate transcription of the heavy chain locus in response to CD4O signaling alone. Experiments are proposed to understand the basis of this functional defect.
StatusFinished
Effective start/end date8/5/017/31/02

Funding

  • National Institute of Allergy and Infectious Diseases: $229,365.00

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