Sterile alpha motif containing 7 (samd7) is a novel crx-regulated transcriptional repressor in the retina.

Inherited retinal diseases are mainly caused by mutations in genes that are highly expressed in photoreceptors of the retina. The majority of these genes is under the control of the transcription factor Cone rod homeobox (Crx), that acts as a master transcription factor in photoreceptors. Using a genome-wide chromatin immunoprecipitation dataset that highlights all potential in vivo targets of Crx, we have identified a novel sterile alpha motif (SAM) domain containing protein, Samd7. mRNA Expression of Samd7 was confined to the late postnatal and adult mouse retina as well as the pineal gland. Using immunohistochemistry and Western blot, we could detect Samd7 protein in the outer nuclear layer of adult mouse retina. Ectopic over-expression in HEK293 cells demonstrated that Samd7 resides in the cytoplasm as well as the nucleus. In vitro electroporation of fluorescent reporters into living mouse retinal cultures revealed that transcription of the Samd7 gene depends on evolutionary conserved Crx motifs located in the first intron enhancer. Moreover, Crx knock-down with shRNA strongly reduced Samd7 reporter activity and endogenous Samd7 protein, indicating that Crx is required for retinal expression of Samd7. Finally, using co-transfections in luciferase reporter assays we found that Samd7 interferes with Crx-dependent transcription. Samd7 suppressed luciferase activity from a reporter plasmid with five Crx consensus repeats in a dose dependent manner and reduced Crx-mediated transactivation of regulatory sequences in the retinoschisin gene and the Samd7 gene itself. Taken together, we have identified a novel retinal SAM domain protein, Samd7, which could act as a transcriptional repressor involved in fine-tuning of Crx-regulated gene expression.

Luteolin triggers global changes in the microglial transcriptome leading to a unique anti-inflammatory and neuroprotective phenotype.

BACKGROUND: Luteolin, a plant derived flavonoid, exerts a variety of pharmacological activities and anti-oxidant properties associated with its capacity to scavenge oxygen and nitrogen species. Luteolin also shows potent anti-inflammatory activities by inhibiting nuclear factor kappa B (NFkB) signaling in immune cells. To better understand the immuno-modulatory effects of this important flavonoid, we performed a genome-wide expression analysis in pro-inflammatory challenged microglia treated with luteolin and conducted a phenotypic and functional characterization. METHODS: Resting and LPS-activated BV-2 microglia were treated with luteolin in various concentrations and mRNA levels of pro-inflammatory markers were determined. DNA microarray experiments and bioinformatic data mining were performed to capture global transcriptomic changes following luteolin stimulation of microglia. Extensive qRT-PCR analyses were carried out for an independent confirmation of newly identified luteolin-regulated transcripts. The activation state of luteolin-treated microglia was assessed by morphological characterization. Microglia-mediated neurotoxicity was assessed by quantifying secreted nitric oxide levels and apoptosis of 661W photoreceptors cultured in microglia-conditioned medium. RESULTS: Luteolin dose-dependently suppressed pro-inflammatory marker expression in LPS-activated microglia and triggered global changes in the microglial transcriptome with more than 50 differentially expressed transcripts. Pro-inflammatory and pro-apoptotic gene expression was effectively blocked by luteolin. In contrast, mRNA levels of genes related to anti-oxidant metabolism, phagocytic uptake, ramification, and chemotaxis were significantly induced. Luteolin treatment had a major effect on microglial morphology leading to ramification of formerly amoeboid cells associated with the formation of long filopodia. When co-incubated with luteolin, LPS-activated microglia showed strongly reduced NO secretion and significantly decreased neurotoxicity on 661W photoreceptor cultures. CONCLUSIONS: Our findings confirm the inhibitory effects of luteolin on pro-inflammatory cytokine expression in microglia. Moreover, our transcriptomic data suggest that this flavonoid is a potent modulator of microglial activation and affects several signaling pathways leading to a unique phenotype with anti-inflammatory, anti-oxidative, and neuroprotective characteristics. With the identification of several novel luteolin-regulated genes, our findings provide a molecular basis to understand the versatile effects of luteolin on microglial homeostasis. The data also suggest that luteolin could be a promising candidate to develop immuno-modulatory and neuroprotective therapies for the treatment of neurodegenerative disorders.