Rajeev Sivasankaran

Phosphorylated FTY720 promotes astrocyte migration through sphingosine-1-phosphate receptors

Sphingosine‐1‐phosphate (S1P) receptors are widely expressed in the central nervous system where they are thought to regulate glia cell function. The phosphorylated version of fingolimod/FTY720 (FTY720P) is active on a broad spectrum of S1P receptors and the parent compound is currently in phase III clinical trials for the treatment of multiple sclerosis. Here, we aimed to identify which cell type(s) and S1P receptor(s) of the central nervous system are targeted by FTY720P. Using calcium imaging in mixed cultures from embryonic rat cortex we show that astrocytes are the major cell type responsive to FTY720P in this assay. In enriched astrocyte cultures, we detect expression of S1P1 and S1P3 receptors and demonstrate that FTY720P activates Gi protein‐mediated signaling cascades. We also show that FTY720P as well as the S1P1‐selective agonist SEW2871 stimulate astrocyte migration. The data indicate that FTY720P exerts its effects on astrocytes predominantly via the activation of S1P1 receptors, whereas S1P signals through both S1P1 and S1P3 receptors. We suggest that this distinct pharmacological profile of FTY720P, compared with S1P, could play a role in the therapeutic effects of FTY720 in multiple sclerosis.

Identification of Novel Genes and Pathways Regulating SREBP Transcriptional Activity

Background Lipid metabolism in mammals is orchestrated by a family of transcription factors called sterol regulatory element-binding proteins (SREBPs) that control the expression of genes required for the uptake and synthesis of cholesterol, fatty acids, and triglycerides. SREBPs are thus essential for insulin-induced lipogenesis and for cellular membrane homeostasis and biogenesis. Although multiple players have been identified that control the expression and activation of SREBPs, gaps remain in our understanding of how SREBPs are coordinated with other physiological pathways. Methodology To identify novel regulators of SREBPs, we performed a genome-wide cDNA over-expression screen to identify proteins that might modulate the transcription of a luciferase gene driven from an SREBP–specific promoter. The results were verified through secondary biological assays and expression data were analyzed by a novel application of the Gene Set Enrichment Analysis (GSEA) method. Conclusions/Significance We screened 10,000 different cDNAs and identified a number of genes and pathways that have previously not been implicated in SREBP control and cellular cholesterol homeostasis. These findings further our understanding of lipid biology and should lead to new insights into lipid associated disorders.

Genetic circuitry of Survival motor neuron, the gene underlying spinal muscular atrophy

Significance Spinal muscular atrophy (SMA), the leading genetic cause of infant mortality, is a devastating neurodegenerative disease caused by reduced levels of Survival Motor Neuron (SMN) gene activity. Despite well-characterized aspects of the involvement of SMN in small nuclear ribonucleoprotein biogenesis, the gene circuitry affecting SMN activity remains obscure. Here, we use Drosophila as a model system to integrate results from large-scale genetic and proteomic studies and bioinformatic analyses to define a unique SMN interactome to provide a basis for a better understanding of SMA. Such efforts not only help dissect Smn biology but also may point to potential clinically relevant targets. The clinical severity of the neurodegenerative disorder spinal muscular atrophy (SMA) is dependent on the levels of functional Survival Motor Neuron (SMN) protein. Consequently, current strategies for developing treatments for SMA generally focus on augmenting SMN levels. To identify additional potential therapeutic avenues and achieve a greater understanding of SMN, we applied in vivo, in vitro, and in silico approaches to identify genetic and biochemical interactors of the Drosophila SMN homolog. We identified more than 300 candidate genes that alter an Smn-dependent phenotype in vivo. Integrating the results from our genetic screens, large-scale protein interaction studies, and bioinformatic analysis, we define a unique interactome for SMN that provides a knowledge base for a better understanding of SMA.

Synthetic transcription elongation factors license transcription across repressive chromatin

Chemical control of transcription Friedreichs ataxia, a devastating neurodegenerative disease with no effective therapy, is caused by an expansion of intronic repeats and hence a reduced expression of the FXN gene. Erwin et al. synthesized a molecule that specifically targets the expanded repressive repeats. This molecule thereby licenses productive transcription elongation and restores FXN expression to normal levels. In the future, similar interventions may be effective in a diverse array of diseases caused by unstable expansions in microsatellite repeats. Science, this issue p. 1617 A synthetic molecule can surmount barriers to transcription elongation at repressive microsatellite repeats that cause Friedreich’s ataxia. The release of paused RNA polymerase II into productive elongation is highly regulated, especially at genes that affect human development and disease. To exert control over this rate-limiting step, we designed sequence-specific synthetic transcription elongation factors (Syn-TEFs). These molecules are composed of programmable DNA-binding ligands flexibly tethered to a small molecule that engages the transcription elongation machinery. By limiting activity to targeted loci, Syn-TEFs convert constituent modules from broad-spectrum inhibitors of transcription into gene-specific stimulators. Here we present Syn-TEF1, a molecule that actively enables transcription across repressive GAA repeats that silence frataxin expression in Friedreich’s ataxia, a terminal neurodegenerative disease with no effective therapy. The modular design of Syn-TEF1 defines a general framework for developing a class of molecules that license transcription elongation at targeted genomic loci.

Corrigendum: SMN2 splice modulators enhance U1-pre-mRNA association and rescue SMA mice.

Nat. Chem. Biol. 11, 511–517 (2015); published online 1 June 2015; corrected online 15 July 2015 and 11 February 2016 In the version of this article originally published online, the schematic for the construct in Figure 4a was incorrect. A corrected figure has been provided in the HTML and PDF versions of the article.