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The Maf-family leucine zipper transcription factor NRL is essential for rod photoreceptor development and functional maintenance in the mammalian retina. Mutations in NRL are associated with human retinopathies, and loss of Nrl in mice leads to a cone-only retina with the complete absence of rods. Among the highly down-regulated genes in the Nrl(-/-) retina, we identified receptor expression enhancing protein 6 (Reep6), which encodes a member of a family of proteins involved in shaping of membrane tubules and transport of G-protein coupled receptors. Here, we demonstrate the expression of a novel Reep6 isoform (termed Reep6.1) in the retina by exon-specific Taqman assay and rapid analysis of complementary deoxyribonucleic acid (cDNA) ends (5'-RACE). The REEP6.1 protein includes 27 additional amino acids encoded by exon 5 and is specifically expressed in rod photoreceptors of developing and mature retina. Chromatin immunoprecipitation assay identified NRL binding within the Reep6 intron 1. Reporter assays in cultured cells and transfections in retinal explants mapped an intronic enhancer sequence that mediated NRL-directed Reep6.1 expression. We also demonstrate that knockdown of Reep6 in mouse and zebrafish resulted in death of retinal cells. Our studies implicate REEP6.1 as a key functional target of NRL-centered transcriptional regulatory network in rod photoreceptors., (Published by Oxford University Press 2014. This work is written by (a) US Government employee(s) and is in the public domain in the US.)

Cone photoreceptors are the primary initiator of visual transduction in the human retina. Dysfunction or death of rod photoreceptors precedes cone loss in many retinal and macular degenerative diseases, suggesting a rod-dependent trophic support for cone survival. Rod differentiation and homeostasis are dependent on the basic motif leucine zipper transcription factor neural retina leucine zipper (NRL). The loss of Nrl (Nrl(-/-)) in mice results in a retina with predominantly S-opsin-containing cones that exhibit molecular and functional characteristics of wild-type cones. Here, we report that Nrl(-/-) retina undergoes a rapid but transient period of degeneration in early adulthood, with cone apoptosis, retinal detachment, alterations in retinal vessel structure, and activation and translocation of retinal microglia. However, cone degeneration stabilizes by 4 months of age, resulting in a thinner but intact outer nuclear layer with residual cones expressing S- and M-opsins and a preserved photopic electroretinogram. At this stage, microglia translocate back to the inner retina and reacquire a quiescent morphology. Gene profiling analysis during the period of transient degeneration reveals misregulation of genes related to stress response and inflammation, implying their involvement in cone death. The Nrl(-/-) mouse illustrates the long-term viability of cones in the absence of rods and retinal pigment epithelium defects in a rodless retina. We propose that Nrl(-/-) retina may serve as a model for elucidating mechanisms of cone homeostasis and degeneration that would be relevant to understanding diseases of the cone-dominant human macula.

Background: The circadian clock of Drosophila is able to drive behavioral rhythms for many weeks in continuous darkness (DD). The endogenous rhythm generator is thought to be generated by interlocked molecular feedback loops involving circadian transcriptional and posttranscriptional regulation of several clock genes, including period. However, all attempts to demonstrate sustained rhythms of clock gene expression in DD have failed, making it difficult to link the molecular clock models with the circadian behavioral rhythms. Here we restricted expression of a novel period-luciferase transgene to certain clock neurons in the Drosophila brain, permitting us to monitor reporter gene activity in these cells in real-time., Results: We show that only a subset of the previously described pacemaker neurons is able to sustain PERIOD protein oscillations after 5 days in constant darkness. In addition, we identified a sustained and autonomous molecular oscillator in a group of clock neurons in the dorsal brain with heretofore unknown function. We found that these "dorsal neurons" (DNs) can synchronize behavioral rhythms and that light input into these cells involves the blue-light photoreceptor cryptochrome., Conclusions: Our results suggest that the DNs play a prominent role in controlling locomotor behavior when flies are exposed to natural light-dark cycles. Analysis of similar "stable mosaic" transgenes should help to reveal the function of the other clock neuronal clusters within the fly brain.