Compound Heterozygous Mutation of Rag1 Leading to Omenn Syndrome

Omenn syndrome is a primary immunodeficiency disorder, featuring susceptibility to infections and autoreactive T cells and resulting from defective genomic rearrangement of genes for the T cell and B cell receptors. The most frequent etiologies are hypomorphic mutations in “non-core” regions of the Rag1 or Rag2 genes, the protein products of which are critical members of the cellular apparatus for V(D)J recombination. In this report, we describe an infant with Omenn syndrome with a previously unreported termination mutation (p.R142*) in Rag1 on one allele and a partially characterized substitution mutation (p.V779M) in a “core” region of the other Rag1 allele. Using a cellular recombination assay, we found that while the p.R142* mutation completely abolished V(D)J recombination activity, the p.V779M mutation conferred a severe, but not total, loss of V(D)J recombination activity. The recombination defect of the V779 mutant was not due to overall misfolding of Rag1, however, as this mutant supported wild-type levels of V(D)J cleavage. These findings provide insight into the role of this poorly understood region of Rag1 and support the role of Rag1 in a post-cleavage stage of recombination.     

Isolation and characterization of a substitution mutation in the ompR gene of Salmonella typhimurium LT2.

The expression of the genes ompC and ompF encoding major outer membrane proteins is dependent on the ompR-envZ operon. Here we describe the isolation and characterization of an ompR mutation, a single-base-pair change, that results in an Arg-to-Cys substitution. When present in multiple copies, the mutant allele conferred a dominant OmpC- OmpF+ phenotype. Furthermore, the mutant allele exhibited allele-specific negative complementation with other ompR mutations. This ability, together with its dominant character, suggested that the OmpR protein is capable of multimerization.     

The Arabidopsis ATRIP ortholog is required for a programmed response to replication inhibitors

The programmed response to replication inhibitors in eukaryotic cells requires the protein kinase ATR (ataxia telangiectasia mutated and rad3-related), which is activated primarily through the persistence of replication protein A (RPA)-bound single-stranded DNA at stalled replication forks and sites of DNA damage undergoing excision repair. Once activated, ATR initiates a cascade of events, including cell-cycle arrest and induction of DNA repair, to mitigate the mutagenic effects of DNA replication in the presence of damage and/or blockage. While many of the molecular regulators of ATR have been determined in yeast and animal cells, little is known about ATR regulation in plants. To genetically define ATR regulatory pathways in Arabidopsis, we describe here a genetic screen for identifying mutants that display a characteristic phenotype of Arabidopsis atr null mutants – hypersensitivity to the replication blocking agent hydroxyurea (HU). Employing this screen, we isolated a novel mutant, termed hus2 (hydroxyurea-sensitive), that displays hypersensitivity to HU, aphidicolin and ionizing radiation, similar to atr mutants. In addition, cell-cycle progression in response to replication blocks and ionizing radiation is defective in hus2 , displaying a nearly identical phenotype to atr mutants. Positional cloning of hus2 reveals a gene sequence similar to yeast Rad26/Ddc2 and ATRIP (ATR interacting protein), suggesting that hus2 encodes an Arabidopsis ATRIP ortholog.