This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Primary immunodeficiency diseases are unique human models to study the molecular events that ultimately lead to an efficient immune response. With the support of this grant, our group has participated in the identification of genes involved in X-linked immunodeficiency diseases and in the functional analysis of the products of these genes. The understanding of the molecular basis for immune disorders has contributed greatly to the concept of receptor-ligand interaction, and the principles of lymphocyte activation, differentiation, signal transduction and immunoglobulin isotype switching.
Three specific aims will be addressed. (1) Elucidation of the gene responsible for the Wiskott-Aldrich syndrome (WAS) and its product, the WAS protein. We have been part of a team that recently isolated the gene mutated in patients with WAS, a disorder in which most hematopoietic cell lineages are affected. Our collection of B and T cell lines derived from members of over 50 affected families will permit a representative mutation analysis of WAS and will answer the question of whether a correlation exists between clinical phenotype and type of mutation. Based on the amino acid composition derived from the known cDNA sequence, we have outlined strategies to analyze the nature of the WAS protein. We will focus on WAS gene expression and its control, its distribution within the cell, the physical and functional properties of the WAS protein, will identify other proteins that interact with WASP. (2) Explore the role of defective transcriptional activation of T cells in patients with common variable immunodeficiency (CVI). We have identified a subgroup of CVI patients who have in common a broad defect of T cell activation, suggesting that the transcription of multiple lymphokines and membrane bound activation molecules may be abnormal. Using gelshift assays, we will determine if the defect is due to abnormal binding of NF-AT or other nuclear factors related to the IL-2 gene. If decreased binding is demonstrated, we will search for defective production of proteins of the NF-AT complex and analyze its distribution in the cytoplasm and in the nuclear fraction, and its phosphorylation state. (3) Explore the feasibility of gene therapy for patients with X-linked immunodeficiencies. We have initiated collaboration with several investigators involved in the design of viral vectors that efficiently transfer the candidate genes into target cells, e.g., T and B cell lines or stem cells derived from patients.
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