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Anand Swaroop - One of the best experts on this subject based on the ideXlab platform.
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the transcription factor Neural Retina leucine zipper nrl controls photoreceptor specific expression of myocyte enhancer factor mef2c from an alternative promoter
Journal of Biological Chemistry, 2011Co-Authors: Hong Hao, Raghuveer Singh Mali, Anand Swaroop, Padmaja Tummala, Eduardo Guzman, Janina Gregorski, Kenneth P MittonAbstract:Neural Retina leucine zipper (NRL) is an essential transcription factor for cell fate specification and functional maintenance of rod photoreceptors in the mammalian Retina. In the Nrl−/− mouse Retina, photoreceptor precursors fail to produce rods and generate functional cone photoreceptors that predominantly express S-opsin. Previous global expression analysis using microarrays revealed dramatically reduced expression of myocyte enhancer factor Mef2c in the adult Nrl−/− Retina. We undertook this study to examine the biological relevance of Mef2c expression in Retinal rod photoreceptors. Bioinformatics analysis, rapid analysis of cDNA ends (5′-RACE), and reverse transcription coupled with qPCR using splice site-specific oligonucleotides suggested that Mef2c is expressed in the mature Retina from an alternative promoter. Chromatin immunoprecipitation (ChIP) studies showed the association of active RNA polymerase II and acetylated histone H3 just upstream of Mef2c exon 4, providing additional evidence for the utilization of an alternative promoter in the Retina. In concordance, we observed the binding of NRL to a putative NRL-response element (NRE) at this location by ChIP-seq and electrophoretic mobility shift assays. NRL also activated the Mef2c alternative promoter in vitro and in vivo. Notably, MEF2C could support Rhodopsin promoter activity in rod photoreceptors. We conclude that Mef2c expression from an alternative promoter in the Retina is regulated by NRL. Our studies also implicate MEF2C as a transcriptional regulator of homeostasis in rod photoreceptor cells.
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combinatorial regulation of photoreceptor differentiation factor Neural Retina leucine zipper gene nrl revealed by in vivo promoter analysis
Journal of Biological Chemistry, 2011Co-Authors: Marieaudrey Ines Kautzmann, Douglas S Kim, Mariepaule Felderschmittbuhl, Anand SwaroopAbstract:Development and homeostasis require stringent spatiotemporal control of gene expression patterns that are established, to a large extent, by combinatorial action of transcription regulatory proteins. The bZIP transcription factor NRL (Neural Retina leucine zipper) is critical for rod versus cone photoreceptor cell fate choice during Retinal development and acts as a molecular switch to produce rods from postmitotic precursors. Loss of Nrl in mouse leads to a cone-only Retina, whereas ectopic expression of Nrl in photoreceptor precursors generates rods. To decipher the transcriptional regulatory mechanisms upstream of Nrl, we identified putative cis-control elements in the Nrl promoter/enhancer region by examining cross-species sequence conservation. Using in vivo transfection of promoter-reporter constructs into the mouse Retina, we show that a 0.9-kb sequence upstream of the Nrl transcription initiation site is sufficient to drive reporter gene expression in photoreceptors. We further define a 0.3-kb sequence including a proximal promoter (cluster A1) and an enhancer (cluster B) that can direct rod-specific expression in vivo. Electrophoretic mobility shift assays using mouse Retinal nuclear extracts, in combination with specific antibodies, demonstrate the binding of retinoid-related orphan nuclear receptor β (RORβ), cone rod homeobox, orthodenticle homolog 2, and cyclic AMP response element-binding protein to predicted consensus elements within clusters A and B. Our studies demonstrate Nrl as a direct transcriptional target of RORβ and suggest that combinatorial action of multiple regulatory factors modulates the expression of Nrl in developing and mature Retina.
Thomas A Reh - One of the best experts on this subject based on the ideXlab platform.
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extraocular mesenchyme patterns the optic vesicle during early eye development in the embryonic chick
Development, 2000Co-Authors: Sabine Fuhrmann, Edward M Levine, Thomas A RehAbstract:The vertebrate eye develops from the neuroepithelium of the ventral forebrain by the evagination and formation of the optic vesicle. Classical embryological studies have shown that the surrounding extraocular tissues - the surface ectoderm and extraocular mesenchyme - are necessary for normal eye growth and differentiation. We have used explant cultures of chick optic vesicles to study the regulation of Retinal pigmented epithelium (RPE) patterning and differentiation during early eye development. Our results show that extraocular mesenchyme is required for the induction and maintenance of expression of the RPE-specific genes Mitf and Wnt13 and the melanosomal matrix protein MMP115. In the absence of extraocular tissues, RPE development did not occur. Replacement of the extraocular mesenchyme with cranial mesenchyme, but not lateral plate mesoderm, could rescue expression of the RPE-marker Mitf. In addition to activating expression of RPE-specific genes, the extraocular mesenchyme inhibits the expression of the Neural Retina-specific transcription factor Chx10 and downregulates the eye-specific transcription factors Pax6 and Optx2. The TGF(β) family member activin can substitute for the extraocular mesenchyme by promoting expression of the RPE-specific genes and downregulating expression of the Neural Retina-specific markers. These data indicate that extraocular mesenchyme, and possibly an activin-like signal, pattern the domains of the optic vesicle into RPE and Neural Retina.
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fibroblast growth factors are necessary for Neural Retina but not pigmented epithelium differentiation in chick embryos
Development, 1997Co-Authors: Catrin Pittack, Gerald B Grunwald, Thomas A RehAbstract:During eye development, optic vesicles evaginate laterally from the Neural tube and develop into two bilayered eye cups that are composed of an outer pigment epithelium layer and an inner Neural Retina layer. Despite their similar embryonic origin, the pigment epithelium and Neural Retina differentiate into two very distinct tissues. Previous studies have demonstrated that the developmental potential of the pigmented epithelial cells is not completely restricted; until embryonic day 4.5 in chick embryos, the cells are able to switch their phenotype and differentiate into Neural Retina when treated with fibroblast growth factors (FGF) (Park, C. M., and Hollenberg, M. J. (1989). Dev. Biol. 134, 201–205; Pittack, C., Jones, M., and Reh, T. A. 1991). Development 113, 577–588; Guillemot, F. and Cepko, C. L. (1992). Development 114, 743–754). These studies motivated us to test whether FGF is necessary for Neural Retina differentiation during the initial stages of eye cup development. Optic vesicles from embryonic day 1.5 chick were cultured for 24 hours as explants in the presence of FGF or neutralizing antibodies to FGF2. The cultured optic vesicles formed eye cups that contained a lens vesicle, Neural Retina and pigmented epithelium, based on morphology and expression of Neural and pigmented epithelium-specific antigens. Addition of FGF to the optic vesicles caused the presumptive pigmented epithelium to undergo neuronal differentiation and, as a consequence, a double Retina was formed. By contrast, neutralizing antibodies to FGF2 blocked Neural differentiation in the presumptive Neural Retina, without affecting pigmented epithelial cell differentiation. These data, along with evidence for expression of several FGF family members and their receptors in the developing eye, indicate that members of the FGF family may be required for establishing the distinction between the Neural Retina and pigmented epithelium in the optic vesicle.
Kenneth P Mitton - One of the best experts on this subject based on the ideXlab platform.
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the transcription factor Neural Retina leucine zipper nrl controls photoreceptor specific expression of myocyte enhancer factor mef2c from an alternative promoter
Journal of Biological Chemistry, 2011Co-Authors: Hong Hao, Raghuveer Singh Mali, Anand Swaroop, Padmaja Tummala, Eduardo Guzman, Janina Gregorski, Kenneth P MittonAbstract:Neural Retina leucine zipper (NRL) is an essential transcription factor for cell fate specification and functional maintenance of rod photoreceptors in the mammalian Retina. In the Nrl−/− mouse Retina, photoreceptor precursors fail to produce rods and generate functional cone photoreceptors that predominantly express S-opsin. Previous global expression analysis using microarrays revealed dramatically reduced expression of myocyte enhancer factor Mef2c in the adult Nrl−/− Retina. We undertook this study to examine the biological relevance of Mef2c expression in Retinal rod photoreceptors. Bioinformatics analysis, rapid analysis of cDNA ends (5′-RACE), and reverse transcription coupled with qPCR using splice site-specific oligonucleotides suggested that Mef2c is expressed in the mature Retina from an alternative promoter. Chromatin immunoprecipitation (ChIP) studies showed the association of active RNA polymerase II and acetylated histone H3 just upstream of Mef2c exon 4, providing additional evidence for the utilization of an alternative promoter in the Retina. In concordance, we observed the binding of NRL to a putative NRL-response element (NRE) at this location by ChIP-seq and electrophoretic mobility shift assays. NRL also activated the Mef2c alternative promoter in vitro and in vivo. Notably, MEF2C could support Rhodopsin promoter activity in rod photoreceptors. We conclude that Mef2c expression from an alternative promoter in the Retina is regulated by NRL. Our studies also implicate MEF2C as a transcriptional regulator of homeostasis in rod photoreceptor cells.
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fiz1 is part of the regulatory protein complex on active photoreceptor specific gene promoters in vivo
BMC Molecular Biology, 2008Co-Authors: Raghuveer Singh Mali, Shiming Chen, Guang Hua Peng, Xiao Zhang, Loan Dang, Kenneth P MittonAbstract:FIZ1 (Flt-3 Interacting Zinc-finger) is a broadly expressed protein of unknown function. We reported previously that in the mammalian Retina, FIZ1 interacts with NRL (Neural-Retina Leucine-zipper), an essential transcriptional activator of rod photoreceptor-specific genes. The concentration of FIZ1 in the Retina increases during photoreceptor terminal maturation, when two key transcription factors NRL and CRX (Cone-Rod Homeobox) become detectable on the promoters of photoreceptor-specific genes (i.e. Rhodopsin, Pde6b). To determine if FIZ1 is involved in regulating CRX-mediated transcriptional activation, we examined FIZ1 subcellular location in mouse Neural Retina, its ability to interact with CRX, and its association with CRX/NRL target genes. FIZ1 is present in the nucleus of adult photoreceptors as well as other Retinal neurons as shown by transmission electron microscopy with nano-gold labeling. FIZ1 and CRX were co-precipitated from Retinal nuclear extracts with antibodies to either protein. Chromatin immunoprecipitation (ChIP) assays revealed that FIZ1 is part of the protein complex on several rod and cone gene promoters, within photoreceptor cells of the mouse Retina. FIZ1 complexes with CRX or NRL on known NRL- and CRX-responsive elements, as shown by electrophoretic mobility shift assays with FIZ1 antibody. FIZ1 can directly bind to CRX, as demonstrated using yeast two-hybrid and GST pull-down assays. Co-transfection assays demonstrated that FIZ1 increases CRX-mediated activation of Opsin test promoters. Quantitative ChIP analysis revealed an increased association of FIZ1 with the Rhodopsin promoter in adult (P-25) Neural Retina versus immature (P-3) Neural Retina. The quantity of transcriptionally active RNA Polymerase-II within the Rhodopsin gene (Rho) was significantly increased in the adult Neural Retina, compared to the immature Retina. FIZ1 directly interacts with CRX to enhance CRX's transactivation activity for target genes. Developmentally, in Neural Retina tissue, the increased association of FIZ1 with CRX target genes corresponds to an increased association of transcriptionally active Pol-II within the Rho gene. Together with previous findings, our results suggest that FIZ1 may act as a transcriptional co-regulator of photoreceptor-specific genes, recruited by at least two photoreceptor-specific transcription factors, CRX and NRL. Further studies are underway to elucidate the exact role of FIZ1 in photoreceptor gene expression, development and maintenance.
Sébastien Augustin - One of the best experts on this subject based on the ideXlab platform.
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Foxg1-Cre mediated Lrp2 inactivation in the developing mouse Neural Retina, ciliary and Retinal pigment epithelia models congenital high myopia
PLoS ONE, 2015Co-Authors: Olivier Cases, Manuel Simonutti, Antoine Joseph, Antoine Obry, Mathieu D. Santin, Sirine Ben-yacoub, Michel Pâques, Sabine Amsellem-levera, Ana Bribian, Sébastien AugustinAbstract:Myopia is a common ocular disorder generally due to increased axial length of the eye-globe. Its extreme form high myopia (HM) is a multifactorial disease leading to Retinal and scleral dam-age, visual impairment or loss and is an important health issue. Mutations in the endocytic receptor LRP2 gene result in Donnai-Barrow (DBS) and Stickler syndromes, both characterized by HM. To clearly establish the link between Lrp2 and congenital HM we inactivated Lrp2 in the mouse forebrain including the Neural Retina and the Retinal and ciliary pigment epithelia. High resolution in vivo MRI imaging and ophthalmological analyses showed that the adult Lrp2-defi-cient eyes were 40% longer than the control ones mainly due to an excessive elongation of the vitreal chamber. They had an apparently normal intraocular pressure and developed chorioret-inal atrophy and posterior scleral staphyloma features reminiscent of human myopic retinopa-thy. Immunomorphological and ultrastructural analyses showed that increased eye lengthening was first observed by post-natal day 5 (P5) and that it was accompanied by a rapid decrease of the bipolar, photoreceptor and Retinal ganglion cells, and eventually the optic nerve axons. It was followed by scleral thinning and collagen fiber disorganization, essentially in the posterior pole. We conclude that the function of LRP2 in the ocular tissues is necessary for normal eye growth and that the Lrp2-deficient eyes provide a unique tool to further study human HM.
Joseph C Corbo - One of the best experts on this subject based on the ideXlab platform.
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transcriptional regulation of Neural Retina leucine zipper nrl a photoreceptor cell fate determinant
Journal of Biological Chemistry, 2011Co-Authors: Cynthia L Montana, Shiming Chen, Karen A Lawrence, Natecia L Williams, Nicholas M Tran, G H Peng, Joseph C CorboAbstract:The transcription factor Neural Retina leucine zipper (Nrl) is a critical determinant of rod photoreceptor cell fate and a key regulator of rod differentiation. Nrl−/− rod precursors fail to turn on rod genes and instead differentiate as cones. Furthermore, NRL mutations in humans cause retinitis pigmentosa. Despite the developmental and clinical significance of this gene, little is known about the transcriptional regulation of Nrl itself. In this study, we sought to define the cis- and trans-acting factors responsible for initiation and maintenance of Nrl transcription in the mouse Retina. Utilizing a quantitative mouse Retinal explant electroporation assay, we discovered a phylogenetically conserved, 30-base pair region immediately upstream of the transcription start site that is required for Nrl promoter activity. This region contains binding sites for the Retinal transcription factors CRX, OTX2, and RORβ, and point mutations in these sites completely abolish promoter activity in living Retinas. Gel-shift experiments show that CRX, OTX2, and RORβ can bind to the critical region in vitro, whereas ChIP experiments demonstrate binding of CRX and OTX2 to the critical region in vivo. Thus, our results indicate that CRX, OTX2, and RORβ directly regulate Nrl transcription by binding to critical sites within the Nrl promoter. We propose a model in which Nrl expression is primarily initiated by OTX2 and RORβ and later maintained at high levels by CRX and RORβ.