A new system for lineage ablation is based on transgenic expression of a diphtheria toxin receptor (DTR) in mouse cells and application of diphtheria toxin (DT). To streamline this approach, we generated Cre-inducible DTR transgenic mice (iDTR) in which Cre-mediated excision of a STOP cassette renders cells sensitive to DT. We tested the iDTR strain by crossing to the T cell- and B cell-specific CD4-Cre and CD19-Cre strains, respectively, and observed efficient ablation of T and B cells after exposure to DT. In MOGi-Cre/iDTR double transgenic mice expressing Cre recombinase in oligodendrocytes, we observed myelin loss after intraperitoneal DT injections. Thus, DT crosses the blood-brain barrier and promotes cell ablation in the central nervous system. Notably, we show that the developing DT-specific antibody response is weak and not neutralizing, and thus does not impede the efficacy of DT. Our results validate the use of iDTR mice as a tool for cell ablation in vivo.

It is generally assumed that new genes would arise by gene duplication mechanisms, because the signals for regulation and transcript processing would be unlikely to evolve in parallel with a new gene function. We have identified here a transcript in the house mouse (Mus musculus) that has arisen within the past 2.5-3.5 million years in a large intergenic region. The region is present in many mammals, including humans, allowing us to exclude the involvement of gene duplication, transposable elements, or other genome rearrangements, which are typically found for other cases of newly evolved genes. The gene has three exons, shows alternative splicing, and is specifically expressed in postmeiotic cells of the testis. The transcript is restricted to species within the genus Mus and its emergence correlates with indel mutations in the 5' regulatory region of the transcript. A recent selective sweep is associated with the transcript region in M. m. musculus populations. A knockout in the laboratory strain BL6 results in reduced sperm motility and reduced testis weight. Our results show that cryptic signals for transcript regulation and processing exist in intergenic regions and can become the basis for the evolution of a new functional gene.

Herein, we report ap re-synthetic pore environment design strategy to achieve stable methyl-functionalized metalorganic frameworks (MOFs) for preferential SO 2 binding and thus enhanced low (partial) pressure SO 2 adsorption and SO 2 / CO 2 separation. The enhanced sorption performance is for the first time attributed to an optimal pore sizeb yi ncreasing methyl group densities at the benzenedicarboxylate linker in [Ni 2 (BDC-X) 2 DABCO] (BDC-X = mono-, di-, and tetramethyl-1,4-benzenedicarboxylate/terephthalate;D ABCO = 1,4-diazabicyclo[2,2,2]octane). Monte Carlo simulations and first-principles density functional theory (DFT) calculations demonstrate the key role of methyl groups within the pore surface on the preferential SO 2 affinity over the parent MOF. The SO 2 separation potential by methyl-functionalized MOFs has been validated by gas sorption isotherms,i deal adsorbed solution theory calculations,s imulated and experimental breakthrough curves,and DFT calculations.

2), it was presumed that anti-IgD Abs targeted resting B cells, and due to a lack of costimulatory molecules, these B cells tolerized MBP-specific T cells. Similarly, it was shown that passive transfer of B cells expressing a myelin peptide prevented the induction of EAE (6 -8) or even EAE relapses (9). One explanation why B cells induce tolerance of naive but not memory T cells might be the need for expression of costimulatory molecules by the APC to activate naive T cells, specifically B7-1 and/or B7-2, but resting B cells do not express these molecules. On the other hand, memory T cells may not need costimulation and could therefore be activated by B cells. A problem of this hypothesis is that also activated B cells, which normally do express B7 molecules, can induce tolerance of T cells (10). It is therefore not clear whether it is indeed the absence of costimulation what causes B cells to induce tolerance.To study the role of B cells in tolerance induction we have generated mice that express an MHC class II-restricted immunodominant T cell epitope of myelin oligodendrocyte glycoprotein (MOG) specifically on B cells. These mice were found to be resistant to EAE induction. We could show that, following interaction of naive T cells with B cells presenting their specific Ag, T cells are partially activated, resulting in very marginal proliferation and up-regulation of coinhibitory molecules such as CTLA-4, B and T lymphocyte attenuator (BTLA), PD-1, and CD5. Subsequent in vivo activation of tolerized T cells leads to their deletion. Thus, we assessed that naive B cells induce peripheral tolerance by inducing expression of negative costimulatory molecules by Ag-specific T cells, followed by Ag-induced cell death (AICD) upon the next Ag encounter. Materials and Methods Generation of invariant chain (Ii) MOG miceThe targeting vector ROSA26STOP*IiMOG was constructed by introduction into the XbaI site of the vector ROSA26 -1 (11); a gift from P. Soriano (Fred Hutchinson Cancer Research Center, Seattle, WA) of a fragment comprising (5Ј to 3Ј): adenoviral splice acceptor, loxP, 2ϫ SV40 polyadenylation signal, FRT-flanked pGK-neo, a STOP cassette (12), loxP, mutant invariant chain (termed IiMOG), and bovine poly(A). The mutant invariant chain (IiMOG) was generated by assembly PCR on Ii template cDNA (derived from plasmid pcEX V3 mIi31, carrying the cDNA of the

While it is established that the functional impact of genetic variation can vary across cell types and states, capturing this diversity remains challenging. Current studies using bulk sequencing either ignore this heterogeneity or use sorted cell populations, reducing discovery and explanatory power. Here, we develop scDALI, a versatile computational framework that integrates information on cellular states with allelic quantifications of single-cell sequencing data to characterize cell-state-specific genetic effects. We apply scDALI to scATAC-seq profiles from developing F1 Drosophila embryos and scRNA-seq from differentiating human iPSCs, uncovering heterogeneous genetic effects in specific lineages, developmental stages, or cell types.