Chimeric IgH-TCRalpha/delta translocations in T lymphocytes mediated by RAG.

Translocations involving the T cell receptor alpha/delta (TCRalpha/delta) chain locus, which bring oncogenes in the proximity of the TCRalpha enhancer, are one of the hallmark features of human T cell malignancies from ataxia telangiectasia (AT) and non-AT patients. These lesions are frequently generated by the fusion of DNA breaks at the TCRalpha/delta locus to a disperse region centromeric of the immunoglobulin heavy chain (IgH) locus. Aberrant VDJ joining accounts for TCRalpha/delta associated DNA cleavage, but the molecular mechanism that leads to generation of the "oncogene partner" DNA break is unclear. Here we show that in ATM deficient primary mouse T cells, IgH/TCRalpha/delta fusions arise at a remarkably similar frequency as in human AT lymphocytes. Recombinase-activating gene (RAG) is responsible for both TCRalpha/delta as well as IgH associated breaks on chromosome 12 (Chr12), which are subject to varying degrees of chromosomal degradation. We suggest a new model for how oncogenic translocations can arise from two non-concerted physiological DSBs.

microRNA-155 regulates the generation of immunoglobulin class-switched plasma cells.

microRNA-155 (miR-155) is expressed by cells of the immune system after activation and has been shown to be required for antibody production after vaccination with attenuated Salmonella. Here we show the intrinsic requirement for miR-155 in B cell responses to thymus-dependent and -independent antigens. B cells lacking miR-155 generated reduced extrafollicular and germinal center responses and failed to produce high-affinity IgG1 antibodies. Gene-expression profiling of activated B cells indicated that miR-155 regulates an array of genes with diverse function, many of which are predicted targets of miR-155. The transcription factor Pu.1 is validated as a direct target of miR155-mediated inhibition. When Pu.1 is overexpressed in wild-type B cells, fewer IgG1 cells are produced, indicating that loss of Pu.1 regulation is a contributing factor to the miR-155-deficient phenotype. Our results implicate post-transcriptional regulation of gene expression for establishing the terminal differentiation program of B cells.

Antisense intergenic transcription in V(D)J recombination.

Antigen receptor genes undergo variable, diversity and joining (V(D)J) recombination, which requires ordered large-scale chromatin remodeling. Here we show that antisense transcription, both genic and intergenic, occurs extensively in the V region of the immunoglobulin heavy chain locus. RNA fluorescence in situ hybridization demonstrates antisense transcription is strictly developmentally regulated and is initiated during the transition from DJ(H) to VDJ(H) recombination and terminates concomitantly with VDJ(H) recombination. Our data show antisense transcription is specific to the V region and suggest transcripts extend across several genes. We propose that antisense transcription remodels the V region to facilitate V(H)-to-DJ(H) recombination. These findings have wider implications for V(D)J recombination of other antigen receptor loci and developmental regulation of multigene loci.

A putative high affinity hexose transporter, hxtA, of Aspergillus nidulans is induced in vegetative hyphae upon starvation and in ascogenous hyphae during cleistothecium formation.

Fungi employ different carbohydrate uptake systems to adapt to certain environmental conditions and to different carbon source concentrations. The hydrolysis of polymeric carbohydrates and the subsequent uptake of monomeric forms may also play a role in development. Aspergillus nidulans accumulates cell wall components during vegetative growth and degrades them during sexual development. We have identified the hxtA (high affinity hexose transporter) gene in a differential library, which was enriched for sexual-specific genes. The hxtA gene is disrupted by 6 introns and predicted to encode a 531 amino acid protein with high similarity to major facilitator superfamily members including the high affinity hexose transporter Gtt1 from Trichoderma harzianum. A. nidulans HxtA contains the 12 predicted transmembrane domains characteristic for this family. Deletion of hxtA did not impair growth of A. nidulans on a variety of carbon sources nor did it inhibit sexual development suggesting redundant sugar uptake systems. We found at least 17 putative hexose transporters in the genome of A. nidulans. Despite the high similarity of HxtA to fungal high affinity glucose transporters, the hxtA gene did not restore growth on glucose of a Saccharomyces cerevisiae mutant, in which all hexose transporters were deleted. Northern blot analysis revealed that the A. nidulans hxtA gene was repressed under high glucose conditions and expressed in vegetative hyphae upon carbon starvation and during sexual development. We found hxtA(p)::sgfp expression in developing cleistothecia specifically in ascogenous hyphae and propose that HxtA is a high affinity glucose transporter involved in sugar metabolism during sexual development.