Temporal lobe epilepsy is a common form of drug-resistant epilepsy that sometimes responds to dietary manipulation such as the 'ketogenic diet'. Here we have investigated the effects of the glycolytic inhibitor 2-deoxy-D-glucose (2DG) in the rat kindling model of temporal lobe epilepsy. We show that 2DG potently reduces the progression of kindling and blocks seizure-induced increases in the expression of brain-derived neurotrophic factor and its receptor, TrkB. This reduced expression is mediated by the transcription factor NRSF, which recruits the NADH-binding co-repressor CtBP to generate a repressive chromatin environment around the BDNF promoter. Our results show that 2DG has anticonvulsant and antiepileptic properties, suggesting that anti-glycolytic compounds may represent a new class of drugs for treating epilepsy. The metabolic regulation of neuronal genes by CtBP will open avenues of therapy for neurological disorders and cancer.
Acquired mutations in the hematopoietic transcription factor GATA binding protein-1 (GATA1) are found in megakaryoblasts from nearly all individuals with Down syndrome with transient myeloproliferative disorder (TMD, also called transient leukemia) and the related acute megakaryoblastic leukemia (DS-AMKL, also called DS-AML M7). These mutations lead to production of a variant GATA1 protein (GATA1s) that is truncated at its N terminus. To understand the biological properties of GATA1s and its relation to DS-AMKL and TMD, we used gene targeting to generate Gata1 alleles that express GATA1s in mice. We show that the dominant action of GATA1s leads to hyperproliferation of a unique, previously unrecognized yolk sac and fetal liver progenitor, which we propose accounts for the transient nature of TMD and the restriction of DS-AMKL to infants. Our observations raise the possibility that the target cells in other leukemias of infancy and early childhood are distinct from those in adult leukemias and underscore the interplay between specific oncoproteins and potential target cells.
Sequencing DNA from several organisms has revealed that duplication and drift of existing genes have primarily molded the contents of a given genome. Though the effect of knocking out or overexpressing a particular gene has been studied in many organisms, no study has systematically explored the effect of adding new links in a biological network. To explore network evolvability, we constructed 598 recombinations of promoters (including regulatory regions) with different transcription or σ-factor genes in Escherichia coli, added over a wild-type genetic background.Here we show that ~95% of new networks are tolerated by the bacteria, that very few alter growth, and that expression level correlates with factor position in the wild-type network hierarchy. Most importantly, we find that certain networks consistently survive over the wild-type under various selection pressures. Therefore new links in the network are rarely a barrier for evolution and can even confer a fitness advantage.The Escherichia coli genome contains ~300 transcription factors (TFs)1,2, organized hierarchically, with few master regulators3-5 (Fig. 1). Only nine regulatory proteins (CRP, FNR, IHF, FIS, ArcA, NarL, H-NS, Fur, and Lrp) control over half of all genes, through direct and indirect interactions6,7. Lower-tier nodes are more sparsely connected and the network structure has a scale-free power-law degree distribution8,9. It has been argued that such networks are particularly robust to random errors since only a few nodes are highlyconnected hubs, whose perturbation would affect the network drastically10. This conclusion is based on the effects of deleting or overexpressing individual nodes. However, the addition of new interactions is thought to be an equally important process for evolution, and the network responses to such changes remain to be systematically explored.Genomes are molded by gene duplication, transfer, mutation and loss. Duplication occurs rapidly in all species11,12 and through mutation serves as material for innovation. This drives cellular network evolution13,14, even though relatively few duplications become fixed in populations11,12. We therefore chose to reconstruct events where an open reading frame (ORF) or gene is duplicated and subsequently becomes linked to a new regulatory input. Thus, promoter region-ORF fusions were constructed on high copy number plasmids Author Information Microarray data are MIAME-compliant and have been deposited at ArrayExpress http://www.ebi.ac.uk/ microarray-as/aer/entry, Accession: E-MEXP-732. Reprints and permissions information is available at npg.nature.com/ reprintsandpermissions. Correspondence and requests for materials should be addressed to M.I. (e-mail: isalan@crg.es and a subset were stably integrated in the E. coli chromosome. Although evolution is unlikely to take such a direct approach, except in rare cases such as gene fusions in chromosomal rearrangements, our approach provides a systematic way to sample the viability of new connectivity. By adding new connection...
Huntington's disease (HD) is a dominantly inherited neurodegenerative disorder caused by expanded CAG repeats in the huntingtin (HTT) gene. Although several palliative treatments are available, there is currently no cure and patients generally die 10-15 y after diagnosis. Several promising approaches for HD therapy are currently in development, including RNAi and antisense analogs. We developed a complementary strategy to test repression of mutant HTT with zinc finger proteins (ZFPs) in an HD model. We tested a "molecular tape measure" approach, using long artificial ZFP chains, designed to bind longer CAG repeats more strongly than shorter repeats. After optimization, stable ZFP expression in a model HD cell line reduced chromosomal expression of the mutant gene at both the protein and mRNA levels (95% and 78% reduction, respectively). This was achieved chromosomally in the context of endogenous mouse HTT genes, with variable CAG-repeat lengths. Shorter wild-type alleles, other genomic CAG-repeat genes, and neighboring genes were unaffected. In vivo, striatal adeno-associated virus viral delivery in R6/2 mice was efficient and revealed dose-dependent repression of mutant HTT in the brain (up to 60%). Furthermore, zinc finger repression was tested at several levels, resulting in protein aggregate reduction, reduced decline in rotarod performance, and alleviation of clasping in R6/2 mice, establishing a proof-of-principle for synthetic transcription factor repressors in the brain.
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