ABSTRACT
A signal initiated by the newly formed Ag receptor is integrated with microenvironmental cues during T cell development to ensure positive selection of CD4+CD8+ progenitors into functionally mature CD4+ or CD8+ T lymphocytes. During this transition, a survival program is initiated, TCR gene recombination ceases, cells migrate into a new thymic microenvironment, the responsiveness of the Ag receptor is tuned, and the cells commit to a specific T lineage. To determine potential regulators of these processes, we used mRNA microarray analysis to compare gene expression changes in CD4+CD8+ thymocytes from TCR transgenic mice that have received a TCR selection signal with those that had not received a signal. We found 129 genes with expression that changed significantly during positive selection, the majority of which were not previously appreciated. A large number of these changes were confirmed by real-time PCR or flow cytometry. We have combined our findings with gene changes reported in the literature to provide a comprehensive report of the genes regulated during positive selection, and we attempted to assign these genes to positive selection process categories.
Subject(s)
Cell Differentiation/genetics , Cell Differentiation/immunology , Gene Expression Profiling , T-Lymphocyte Subsets/cytology , T-Lymphocyte Subsets/metabolism , Thymus Gland/cytology , Thymus Gland/metabolism , Animals , Cell Adhesion/genetics , Cell Adhesion/immunology , Cell Death/genetics , Cell Death/immunology , Cell Lineage/genetics , Cell Lineage/immunology , Cell Movement/genetics , Cell Movement/immunology , Cell Survival/genetics , Cell Survival/immunology , Gene Expression Profiling/methods , Gene Rearrangement, T-Lymphocyte , Kinetics , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Oligonucleotide Array Sequence Analysis/methods , Receptors, Antigen, T-Cell/biosynthesis , Receptors, Antigen, T-Cell/genetics , Receptors, Antigen, T-Cell/metabolism , Recombination, Genetic/immunology , Reverse Transcriptase Polymerase Chain Reaction , T-Lymphocyte Subsets/immunology , Thymus Gland/immunology , Transcription Factors/biosynthesis , Transcription Factors/geneticsABSTRACT
Systemic pseudohypoaldosteronism type I (PHAI) is an autosomal recessive disorder that arises from loss of function mutations of the alpha, beta, or gamma subunit of Epithelial Na(+) Channel (ENaC). In addition to a severe renal phenotype in the neonatal period, patients with PHAI develop a childhood pulmonary syndrome characterized by cough and frequent respiratory infections. We tested a patient, born to consanguineous parents, who presented with dehydration, metabolic acidosis, hyperkalemia, elevated renin and aldosterone levels at birth, and recurrent respiratory symptoms in his first year. He demonstrated defective epithelial Na(+) transport in multiple organs (raised sweat Cl(-), 120 mM; raised salivary Na(+) and Cl(-), 118 and 111 mM, respectively; and little nasal amiloride-sensitive potential difference). No deleterious mutation was identified in the coding region of the three ENaC subunits. Reverse transcriptase-polymerase chain reaction of nasal epithelial RNA showed reduced betaENaC expression, and inability to amplify promoter elements indicated the possibility of a deletion in the 5' region. Using a probe that corresponded to exon 1A of betaENaC, we confirmed a large deletion (> 1,300 bp). In summary, a homozygous mutation in the promoter region of betaENaC leads to PHAI, the first description of a mutation in the regulatory regions of an ENaC subunit leading to a clinical phenotype.
