Publications by authors named "Jerome E Roger"

22 Publications

  • Page 1 of 1

Glycogen Synthase Kinase 3 Regulates the Genesis of Displaced Retinal Ganglion Cells3.

eNeuro 2021 Sep-Oct;8(5). Epub 2021 Oct 6.

Paris-Saclay Institute of Neuroscience, CERTO-Retina France, CNRS, Université Paris-Saclay, 91400, Saclay, France

Glycogen synthase kinase 3 (GSK3) proteins (GSK3α and GSK3β) are key mediators of signaling pathways, with crucial roles in coordinating fundamental biological processes during neural development. Here we show that the complete loss of GSK3 signaling in mouse retinal progenitors leads to microphthalmia with broad morphologic defects. A single wild-type allele of either or is able to rescue this phenotype. In this genetic context, all cell types are present in a functional retina. However, we unexpectedly detected a large number of cells in the inner nuclear layer expressing retinal ganglion cell (RGC)-specific markers (called displaced RGCs, dRGCs) when at least one allele of is expressed. The excess of dRGCs leads to an increased number of axons projecting into the ipsilateral medial terminal nucleus, an area of the brain belonging to the non-image-forming visual circuit and poorly targeted by RGCs in wild-type retina. Transcriptome analysis and optomotor response assay suggest that at least a subset of dRGCs in mutant mice are direction-selective RGCs. Our study thus uncovers a unique role of GSK3 in controlling the production of ganglion cells in the inner nuclear layer, which correspond to dRGCs, a rare and poorly characterized retinal cell type.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1523/ENEURO.0171-21.2021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8496207PMC
October 2021

Diminished retinal complex lipid synthesis and impaired fatty acid β-oxidation associated with human diabetic retinopathy.

JCI Insight 2021 Oct 8;6(19). Epub 2021 Oct 8.

Department of Internal Medicine-Nephrology, University of Michigan, Ann Arbor, Michigan, USA.

BACKGROUNDThis study systematically investigated circulating and retinal tissue lipid determinants of human diabetic retinopathy (DR) to identify underlying lipid alterations associated with severity of DR.METHODSRetinal tissues were retrieved from postmortem human eyes, including 19 individuals without diabetes, 20 with diabetes but without DR, and 20 with diabetes and DR, for lipidomic study. In a parallel study, serum samples from 28 American Indians with type 2 diabetes from the Gila River Indian Community, including 12 without DR, 7 with mild nonproliferative DR (NPDR), and 9 with moderate NPDR, were selected. A mass-spectrometry-based lipidomic platform was used to measure serum and tissue lipids.RESULTSIn the postmortem retinas, we found a graded decrease of long-chain acylcarnitines and longer-chain fatty acid ester of hydroxyl fatty acids, diacylglycerols, triacylglycerols, phosphatidylcholines, and ceramide(NS) in central retina from individuals with no diabetes to those with diabetes with DR. The American Indians' sera also exhibited a graded decrease in circulating long-chain acylcarnitines and a graded increase in the intermediate-length saturated and monounsaturated triacylglycerols from no DR to moderate NPDR.CONCLUSIONThese findings suggest diminished synthesis of complex lipids and impaired mitochondrial β-oxidation of fatty acids in retinal DR, with parallel changes in circulating lipids.TRIAL REGISTRATIONClinicalTrials.gov NCT00340678.FUNDINGThis work was supported by NIH grants R24 DK082841, K08DK106523, R03DK121941, P30DK089503, P30DK081943, P30DK020572, P30 EY007003; The Thomas Beatson Foundation; and JDRF Center for Excellence (5-COE-2019-861-S-B).
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1172/jci.insight.152109DOI Listing
October 2021

Reproducing diabetic retinopathy features using newly developed human induced-pluripotent stem cell-derived retinal Müller glial cells.

Glia 2021 Jul 8;69(7):1679-1693. Epub 2021 Mar 8.

Institut de la Vision, Sorbonne Université, INSERM, CNRS, Paris, France.

Muller glial cells (MGCs) are responsible for the homeostatic and metabolic support of the retina. Despite the importance of MGCs in retinal disorders, reliable and accessible human cell sources to be used to model MGC-associated diseases are lacking. Although primary human MGCs (pMGCs) can be purified from post-mortem retinal tissues, the donor scarcity limits their use. To overcome this problem, we developed a protocol to generate and bank human induced pluripotent stem cell-derived MGCs (hiMGCs). Using a transcriptome analysis, we showed that the three genetically independent hiMGCs generated were homogeneous and showed phenotypic characteristics and transcriptomic profile of pMGCs. These cells expressed key MGC markers, including Vimentin, CLU, DKK3, SOX9, SOX2, S100A16, ITGB1, and CD44 and could be cultured up to passage 8. Under our culture conditions, hiMGCs and pMGCs expressed low transcript levels of RLPB1, AQP4, KCNJ1, KCJN10, and SLC1A3. Using a disease modeling approach, we showed that hiMGCs could be used to model the features of diabetic retinopathy (DR)-associated dyslipidemia. Indeed, palmitate, a major free fatty acid with elevated plasma levels in diabetic patients, induced the expression of inflammatory cytokines found in the ocular fluid of DR patients such as CXCL8 (IL-8) and ANGPTL4. Moreover, the analysis of palmitate-treated hiMGC secretome showed an upregulation of proangiogenic factors strongly related to DR, including ANG2, Endoglin, IL-1β, CXCL8, MMP-9, PDGF-AA, and VEGF. Thus, hiMGCs could be an alternative to pMGCs and an extremely valuable tool to help to understand and model glial cell involvement in retinal disorders, including DR.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/glia.23983DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8252429PMC
July 2021

Altered visual processing in the mdx52 mouse model of Duchenne muscular dystrophy.

Neurobiol Dis 2021 05 5;152:105288. Epub 2021 Feb 5.

Université Paris-Saclay, CNRS, Institut des Neurosciences Paris Saclay, 91190 Gif-sur-Yvette, France. Electronic address:

The mdx52 mouse model of Duchenne muscular dystrophy (DMD) is lacking exon 52 of the DMD gene that is located in a hotspot mutation region causing cognitive deficits and retinal anomalies in DMD patients. This deletion leads to the loss of the dystrophin proteins, Dp427, Dp260 and Dp140, while Dp71 is preserved. The flash electroretinogram (ERG) in mdx52 mice was previously characterized by delayed dark-adapted b-waves. A detailed description of functional ERG changes and visual performances in mdx52 mice is, however, lacking. Here an extensive full-field ERG repertoire was applied in mdx52 mice and WT littermates to analyze retinal physiology in scotopic, mesopic and photopic conditions in response to flash, sawtooth and/or sinusoidal stimuli. Behavioral contrast sensitivity was assessed using quantitative optomotor response (OMR) to sinusoidally modulated luminance gratings at 100% or 50% contrast. The mdx52 mice exhibited reduced amplitudes and delayed implicit times in dark-adapted ERG flash responses, particularly in their b-wave and oscillatory potentials, and diminished amplitudes of light-adapted flash ERGs. ERG responses to sawtooth stimuli were also diminished and delayed for both mesopic and photopic conditions in mdx52 mice and the first harmonic amplitudes to photopic sine-wave stimuli were smaller at all temporal frequencies. OMR indices were comparable between genotypes at 100% contrast but significantly reduced in mdx52 mice at 50% contrast. The complex ERG alterations and disturbed contrast vision in mdx52 mice include features observed in DMD patients and suggest altered photoreceptor-to-bipolar cell transmission possibly affecting contrast sensitivity. The mdx52 mouse is a relevant model to appraise the roles of retinal dystrophins and for preclinical studies related to DMD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.nbd.2021.105288DOI Listing
May 2021

HDAC inhibition ameliorates cone survival in retinitis pigmentosa mice.

Cell Death Differ 2021 04 6;28(4):1317-1332. Epub 2020 Nov 6.

Institute for Ophthalmic Research, Eberhard Karls Universität Tübingen, Tübingen, Germany.

Cone photoreceptor cell death in inherited retinal diseases, such as Retinitis Pigmentosa (RP), leads to the loss of high acuity and color vision and, ultimately to blindness. In RP, a vast number of mutations perturb the structure and function of rod photoreceptors, while cones remain initially unaffected. Extensive rod loss in advanced stages of the disease triggers cone death by a mechanism that is still largely unknown. Here, we show that secondary cone cell death in animal models for RP is associated with increased activity of histone deacetylates (HDACs). A single intravitreal injection of an HDAC inhibitor at late stages of the disease, when the majority of rods have already degenerated, was sufficient to delay cone death and support long-term cone survival in two mouse models for RP, affected by mutations in the phosphodiesterase 6b gene. Moreover, the surviving cones remained light-sensitive, leading to an improvement in visual function. RNA-seq analysis of protected cones demonstrated that HDAC inhibition initiated multi-level protection via regulation of different pro-survival pathways, including MAPK, PI3K-Akt, and autophagy. This study suggests a unique opportunity for targeted pharmacological protection of secondary dying cones by HDAC inhibition and creates hope to maintain vision in RP patients even in advanced disease stages.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41418-020-00653-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8026998PMC
April 2021

The 10q26 Risk Haplotype of Age-Related Macular Degeneration Aggravates Subretinal Inflammation by Impairing Monocyte Elimination.

Immunity 2020 08;53(2):429-441.e8

Sorbonne Université, INSERM, CNRS, Institut de la Vision, 17 rue Moreau, F-75012 Paris, France. Electronic address:

A minor haplotype of the 10q26 locus conveys the strongest genetic risk for age-related macular degeneration (AMD). Here, we examined the mechanisms underlying this susceptibility. We found that monocytes from homozygous carriers of the 10q26 AMD-risk haplotype expressed high amounts of the serine peptidase HTRA1, and HTRA1 located to mononuclear phagocytes (MPs) in eyes of non-carriers with AMD. HTRA1 induced the persistence of monocytes in the subretinal space and exacerbated pathogenic inflammation by hydrolyzing thrombospondin 1 (TSP1), which separated the two CD47-binding sites within TSP1 that are necessary for efficient CD47 activation. This HTRA1-induced inhibition of CD47 signaling induced the expression of pro-inflammatory osteopontin (OPN). OPN expression increased in early monocyte-derived macrophages in 10q26 risk carriers. In models of subretinal inflammation and AMD, OPN deletion or pharmacological inhibition reversed HTRA1-induced pathogenic MP persistence. Our findings argue for the therapeutic potential of CD47 agonists and OPN inhibitors for the treatment of AMD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.immuni.2020.07.021DOI Listing
August 2020

Yap haploinsufficiency leads to Müller cell dysfunction and late-onset cone dystrophy.

Cell Death Dis 2020 08 14;11(8):631. Epub 2020 Aug 14.

Paris-Saclay Institute of Neuroscience, CERTO-Retina France, CNRS, Université Paris-Saclay, Orsay, 91405, France.

Hippo signalling regulates eye growth during embryogenesis through its effectors YAP and TAZ. Taking advantage of a Yap heterozygous mouse line, we here sought to examine its function in adult neural retina, where YAP expression is restricted to Müller glia. We first discovered an unexpected temporal dynamic of gene compensation. At postnatal stages, Taz upregulation occurs, leading to a gain of function-like phenotype characterised by EGFR signalling potentiation and delayed cell-cycle exit of retinal progenitors. In contrast, Yap adult retinas no longer exhibit TAZ-dependent dosage compensation. In this context, Yap haploinsufficiency in aged individuals results in Müller glia dysfunction, late-onset cone degeneration, and reduced cone-mediated visual response. Alteration of glial homeostasis and altered patterns of cone opsins were also observed in Müller cell-specific conditional Yap-knockout aged mice. Together, this study highlights a novel YAP function in Müller cells for the maintenance of retinal tissue homeostasis and the preservation of cone integrity. It also suggests that YAP haploinsufficiency should be considered and explored as a cause of cone dystrophies in human.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41419-020-02860-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7429854PMC
August 2020

Linking YAP to Müller Glia Quiescence Exit in the Degenerative Retina.

Cell Rep 2019 05;27(6):1712-1725.e6

Paris-Saclay Institute of Neuroscience, CERTO-Retina France, CNRS, Univ Paris Sud, Université Paris-Saclay, Orsay 91405, France. Electronic address:

Contrasting with fish or amphibian, retinal regeneration from Müller glia is largely limited in mammals. In our quest toward the identification of molecular cues that may boost their stemness potential, we investigated the involvement of the Hippo pathway effector YAP (Yes-associated protein), which is upregulated in Müller cells following retinal injury. Conditional Yap deletion in mouse Müller cells prevents cell-cycle gene upregulation that normally accompanies reactive gliosis upon photoreceptor cell death. We further show that, in Xenopus, a species endowed with efficient regenerative capacity, YAP is required for their injury-dependent proliferative response. In the mouse retina, where Müller cells do not spontaneously proliferate, YAP overactivation is sufficient to induce their reprogramming into highly proliferative cells. Overall, we unravel a pivotal role for YAP in tuning Müller cell proliferative response to injury and highlight a YAP-EGFR (epidermal growth factor receptor) axis by which Müller cells exit their quiescence state, a critical step toward regeneration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.celrep.2019.04.045DOI Listing
May 2019

Cone-rod homeobox CRX controls presynaptic active zone formation in photoreceptors of mammalian retina.

Hum Mol Genet 2018 10;27(20):3555-3567

Neurobiology-Neurodegeneration & Repair Laboratory, National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USA.

In the mammalian retina, rod and cone photoreceptors transmit the visual information to bipolar neurons through highly specialized ribbon synapses. We have limited understanding of regulatory pathways that guide morphogenesis and organization of photoreceptor presynaptic architecture in the developing retina. While neural retina leucine zipper (NRL) transcription factor determines rod cell fate and function, cone-rod homeobox (CRX) controls the expression of both rod- and cone-specific genes and is critical for terminal differentiation of photoreceptors. A comprehensive immunohistochemical evaluation of Crx-/- (null), CrxRip/+ and CrxRip/Rip (models of dominant congenital blindness) mouse retinas revealed abnormal photoreceptor synapses, with atypical ribbon shape, number and length. Integrated analysis of retinal transcriptomes of Crx-mutants with CRX- and NRL-ChIP-Seq data identified a subset of differentially expressed CRX target genes that encode presynaptic proteins associated with the cytomatrix active zone (CAZ) and synaptic vesicles. Immunohistochemistry of Crx-mutant retina validated aberrant expression of REEP6, PSD95, MPP4, UNC119, UNC13, RGS7 and RGS11, with some reduction in Ribeye and no significant change in immunostaining of RIMS1, RIMS2, Bassoon and Pikachurin. Our studies demonstrate that CRX controls the establishment of CAZ and anchoring of ribbons, but not the formation of ribbon itself, in photoreceptor presynaptic terminals.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/hmg/ddy272DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6454433PMC
October 2018

Repopulating retinal microglia restore endogenous organization and function under CX3CL1-CX3CR1 regulation.

Sci Adv 2018 03 21;4(3):eaap8492. Epub 2018 Mar 21.

Unit on Neuron-Glia Interactions in Retinal Disease, National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USA.

Microglia have been discovered to undergo repopulation following ablation. However, the functionality of repopulated microglia and the mechanisms regulating microglia repopulation are unknown. We examined microglial homeostasis in the adult mouse retina, a specialized neural compartment containing regular arrays of microglia in discrete synaptic laminae that can be directly visualized. Using in vivo imaging and cell-fate mapping techniques, we discovered that repopulation originated from residual microglia proliferating in the central inner retina that subsequently spread by centrifugal migration to fully recapitulate pre-existing microglial distributions and morphologies. Repopulating cells fully restored microglial functions including constitutive "surveying" process movements, behavioral and physiological responses to retinal injury, and maintenance of synaptic structure and function. Microglial repopulation was regulated by CX3CL1-CX3CR1 signaling, slowing in CX3CR1 deficiency and accelerating with exogenous CX3CL1 administration. Microglial homeostasis following perturbation can fully recover microglial organization and function under the regulation of chemokine signaling between neurons and microglia.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1126/sciadv.aap8492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5943055PMC
March 2018

Pias3 is necessary for dorso-ventral patterning and visual response of retinal cones but is not required for rod photoreceptor differentiation.

Biol Open 2017 Jun 15;6(6):881-890. Epub 2017 Jun 15.

Neurobiology-Neurodegeneration and Repair Laboratory, National Eye Institute, National Institutes of Health, 6 Center Drive, Bethesda, MD 20892, USA

Protein inhibitor of activated Stat 3 (Pias3) is implicated in guiding specification of rod and cone photoreceptors through post-translational modification of key retinal transcription factors. To investigate its role during retinal development, we deleted exon 2-5 of the mouse gene, which resulted in complete loss of the Pias3 protein. mice did not show any overt phenotype, and retinal lamination appeared normal even at 18 months. We detected reduced photopic b-wave amplitude by electroretinography following green light stimulation of postnatal day (P)21 retina, suggesting a compromised visual response of medium wavelength (M) cones. No change was evident in response of short wavelength (S) cones or rod photoreceptors until 7 months. Increased S-opsin expression in the M-cone dominant dorsal retina suggested altered distribution of cone photoreceptors. Transcriptome profiling of P21 and 18-month-old retina revealed aberrant expression of a subset of photoreceptor genes. Our studies demonstrate functional redundancy in SUMOylation-associated transcriptional control mechanisms and identify a specific, though limited, role of Pias3 in modulating spatial patterning and optimal function of cone photoreceptor subtypes in the mouse retina.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1242/bio.024679DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5483026PMC
June 2017

Retinal Degeneration Triggers the Activation of YAP/TEAD in Reactive Müller Cells.

Invest Ophthalmol Vis Sci 2017 04;58(4):1941-1953

Paris-Saclay Institute of Neuroscience, CNRS, Univ Paris-Sud, Université Paris-Saclay, Orsay, France 2Centre d'Etude et de Recherche Thérapeutique en Ophtalmologie, Retina France, Orsay, France.

Purpose: During retinal degeneration, Müller glia cells respond to photoreceptor loss by undergoing reactive gliosis, with both detrimental and beneficial effects. Increasing our knowledge of the complex molecular response of Müller cells to retinal degeneration is thus essential for the development of new therapeutic strategies. The purpose of this work was to identify new factors involved in Müller cell response to photoreceptor cell death.

Methods: Whole transcriptome sequencing was performed from wild-type and degenerating rd10 mouse retinas at P30. The changes in mRNA abundance for several differentially expressed genes were assessed by quantitative RT-PCR (RT-qPCR). Protein expression level and retinal cellular localization were determined by western blot and immunohistochemistry, respectively.

Results: Pathway-level analysis from whole transcriptomic data revealed the Hippo/YAP pathway as one of the main signaling pathways altered in response to photoreceptor degeneration in rd10 retinas. We found that downstream effectors of this pathway, YAP and TEAD1, are specifically expressed in Müller cells and that their expression, at both the mRNA and protein levels, is increased in rd10 reactive Müller glia after the onset of photoreceptor degeneration. The expression of Ctgf and Cyr61, two target genes of the transcriptional YAP/TEAD complex, is also upregulated following photoreceptor loss.

Conclusions: This work reveals for the first time that YAP and TEAD1, key downstream effectors of the Hippo pathway, are specifically expressed in Müller cells. We also uncovered a deregulation of the expression and activity of Hippo/YAP pathway components in reactive Müller cells under pathologic conditions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1167/iovs.16-21366DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6024660PMC
April 2017

Requirement for Microglia for the Maintenance of Synaptic Function and Integrity in the Mature Retina.

J Neurosci 2016 Mar;36(9):2827-42

Unit on Neuron-Glia Interactions in Retinal Disease,

Unlabelled: Microglia, the principal resident immune cell of the CNS, exert significant influence on neurons during development and in pathological situations. However, if and how microglia contribute to normal neuronal function in the mature uninjured CNS is not well understood. We used the model of the adult mouse retina, a part of the CNS amenable to structural and functional analysis, to investigate the constitutive role of microglia by depleting microglia from the retina in a sustained manner using genetic methods. We discovered that microglia are not acutely required for the maintenance of adult retinal architecture, the survival of retinal neurons, or the laminar organization of their dendritic and axonal compartments. However, sustained microglial depletion results in the degeneration of photoreceptor synapses in the outer plexiform layer, leading to a progressive functional deterioration in retinal light responses. Our results demonstrate that microglia are constitutively required for the maintenance of synaptic structure in the adult retina and for synaptic transmission underlying normal visual function. Our findings on constitutive microglial function are relevant in understanding microglial contributions to pathology and in the consideration of therapeutic interventions that reduce or perturb constitutive microglial function.

Significance Statement: Microglia, the principal resident immune cell population in the CNS, has been implicated in diseases in the brain and retina. However, how they contribute to the everyday function of the CNS is unclear. Using the model of the adult mouse retina, we examined the constitutive role of microglia by depleting microglia from the retina. We found that in the absence of microglia, retinal neurons did not undergo overt cell death or become structurally disorganized in their processes. However, connections between neurons called synapses begin to break down, leading to a decreased ability of the retina to transmit light responses. Our results indicate that retinal microglia contribute constitutively to the maintenance of synapses underlying healthy vision.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1523/JNEUROSCI.3575-15.2016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4879218PMC
March 2016

Müller glial cell-dependent regeneration of the neural retina: An overview across vertebrate model systems.

Dev Dyn 2016 07 8;245(7):727-38. Epub 2016 Jan 8.

Paris-Saclay Institute of Neuroscience, CNRS, Univ Paris Sud, Université Paris-Saclay, Orsay, France.

Retinal dystrophies are a major cause of blindness for which there are currently no curative treatments. Transplantation of stem cell-derived neuronal progenitors to replace lost cells has been widely investigated as a therapeutic option. Another promising strategy would be to trigger self-repair mechanisms in patients, through the recruitment of endogenous cells with stemness properties. Accumulating evidence in the past 15 year0s has revealed that several retinal cell types possess neurogenic potential, thus opening new avenues for regenerative medicine. Among them, Müller glial cells have been shown to be able to undergo a reprogramming process to re-acquire a stem/progenitor state, allowing them to proliferate and generate new neurons for repair following retinal damages. Although Müller cell-dependent spontaneous regeneration is remarkable in some species such as the fish, it is extremely limited and ineffective in mammals. Understanding the cellular events and molecular mechanisms underlying Müller cell activities in species endowed with regenerative capacities could provide knowledge to unlock the restricted potential of their mammalian counterparts. In this context, the present review provides an overview of Müller cell responses to injury across vertebrate model systems and summarizes recent advances in this rapidly evolving field. Developmental Dynamics 245:727-738, 2016. © 2015 The Authors. Developmental Dynamics published by Wiley Periodicals, Inc.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/dvdy.24375DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4900950PMC
July 2016

Involvement of Bcl-2-associated transcription factor 1 in the differentiation of early-born retinal cells.

J Neurosci 2014 Jan;34(4):1530-41

Institut de la Vision, INSERM UMR_S968, CNRS UMR 7210, Sorbonne Universités, UPMC Univ Paris 06, 75012 Paris, France, Centre Hospitalier National d'Ophtalmologie des Quinze-Vingts, INSERM-DHOS CIC 503, 75 571 PARIS Cedex 12, France, CNRS-UMR 3347, INSERM U1021, Université Paris-sud11, Centre Universitaire, 91405 Orsay, France, and Department of Pharmacology and Toxicology, University of Toronto, Toronto, Ontario M5S 1A8, Canada.

Retinal progenitor proliferation and differentiation are tightly controlled by extrinsic cues and distinctive combinations of transcription factors leading to the generation of retinal cell type diversity. In this context, we have characterized Bcl-2-associated transcription factor (Bclaf1) during rodent retinogenesis. Bclaf1 expression is restricted to early-born cell types, such as ganglion, amacrine, and horizontal cells. Analysis of developing retinas in Bclaf1-deficient mice revealed a reduction in the numbers of retinal ganglion cells, amacrine cells and horizontal cells and an increase in the numbers of cone photoreceptor precursors. Silencing of Bclaf1expression by in vitro electroporation of shRNA in embryonic retina confirmed that Bclaf1 serves to promote amacrine and horizontal cell differentiation. Misexpression of Bclaf1 in late retinal progenitors was not sufficient to directly induce the generation of amacrine and horizontal cells. Domain deletion analysis indicated that the N-terminal domain of Bclaf1 containing an arginine-serine-rich and a bZip domain is required for its effects on retinal cell differentiation. In addition, analysis revealed that Bclaf1 function occurs independently of its interaction with endogenous Bcl-2-related proteins. Altogether, our data demonstrates that Bclaf1expression in postmitotic early-born cells facilitates the differentiation of early retinal precursors into retinal ganglion cells, amacrine cells, and horizontal cells rather than into cone photoreceptors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1523/JNEUROSCI.3227-13.2014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6705315PMC
January 2014

OTX2 loss causes rod differentiation defect in CRX-associated congenital blindness.

J Clin Invest 2014 Feb 2;124(2):631-43. Epub 2014 Jan 2.

Leber congenital amaurosis (LCA) encompasses a set of early-onset blinding diseases that are characterized by vision loss, involuntary eye movement, and nonrecordable electroretinogram (ERG). At least 19 genes are associated with LCA, which is typically recessive; however, mutations in homeodomain transcription factor CRX lead to an autosomal dominant form of LCA. The mechanism of CRX-associated LCA is not understood. Here, we identified a spontaneous mouse mutant with a frameshift mutation in Crx (CrxRip). We determined that CrxRip is a dominant mutation that results in congenital blindness with nonrecordable response by ERG and arrested photoreceptor differentiation with no associated degeneration. Expression of LCA-associated dominant CRX frameshift mutations in mouse retina mimicked the CrxRip phenotype, which was rescued by overexpression of WT CRX. Whole-transcriptome profiling using deep RNA sequencing revealed progressive and complete loss of rod differentiation factor NRL in CrxRip retinas. Expression of NRL partially restored rod development in CrxRip/+ mice. We show that the binding of homeobox transcription factor OTX2 at the Nrl promoter was obliterated in CrxRip mice and ectopic expression of OTX2 rescued the rod differentiation defect. Together, our data indicate that OTX2 maintains Nrl expression in developing rods to consolidate rod fate. Our studies provide insights into CRX mutation-associated congenital blindness and should assist in therapeutic design.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1172/JCI72722DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3904630PMC
February 2014

Determination of posttranslational modifications of photoreceptor differentiation factor NRL: focus on SUMOylation.

Methods Mol Biol 2012 ;884:353-61

Neurobiology Neurodegeneration and Repair Laboratory, National Eye Institute, National Institutes of Health, Bethesda, MD, USA.

Conjugation of SUMO (small ubiquitin-related modifier 1) is a critical posttranslational modification, with significant impact on protein function/activity. Here, we describe direct SUMOylation of GST (glutathione S-transferase)-fusion protein and immunoprecipitation assays for investigating SUMOylation of any protein of interest. We have employed these methods to examine SUMOylation of the basic-motif leucine zipper transcription factor NRL.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-61779-848-1_25DOI Listing
October 2012

Preservation of cone photoreceptors after a rapid yet transient degeneration and remodeling in cone-only Nrl-/- mouse retina.

J Neurosci 2012 Jan;32(2):528-41

Neurobiology-Neurodegeneration and Repair Laboratory, National Eye Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.

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.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1523/JNEUROSCI.3591-11.2012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3567450PMC
January 2012

Next-generation sequencing facilitates quantitative analysis of wild-type and Nrl(-/-) retinal transcriptomes.

Mol Vis 2011 23;17:3034-54. Epub 2011 Nov 23.

Neurobiology-Neurodegeneration and Repair Laboratory, National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USA.

Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived retinal transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT-PCR) methods and to evaluate protocols for optimal high-throughput data analysis.

Methods: Retinal mRNA profiles of 21-day-old wild-type (WT) and neural retina leucine zipper knockout (Nrl(-/-)) mice were generated by deep sequencing, in triplicate, using Illumina GAIIx. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows-Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT-PCR validation was performed using TaqMan and SYBR Green assays.

Results: Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the mouse genome (build mm9) and identified 16,014 transcripts in the retinas of WT and Nrl(-/-) mice with BWA workflow and 34,115 transcripts with TopHat workflow. RNA-seq data confirmed stable expression of 25 known housekeeping genes, and 12 of these were validated with qRT-PCR. RNA-seq data had a linear relationship with qRT-PCR for more than four orders of magnitude and a goodness of fit (R(2)) of 0.8798. Approximately 10% of the transcripts showed differential expression between the WT and Nrl(-/-) retina, with a fold change ≥1.5 and p value <0.05. Altered expression of 25 genes was confirmed with qRT-PCR, demonstrating the high degree of sensitivity of the RNA-seq method. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to retinal function. Data analysis with BWA and TopHat workflows revealed a significant overlap yet provided complementary insights in transcriptome profiling.

Conclusions: Our study represents the first detailed analysis of retinal transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3233386PMC
April 2012

Excess cones in the retinal degeneration rd7 mouse, caused by the loss of function of orphan nuclear receptor Nr2e3, originate from early-born photoreceptor precursors.

Hum Mol Genet 2011 Nov 3;20(21):4102-15. Epub 2011 Aug 3.

Department of Ophthalmology and Visual Sciences, University of Michigan, Ann Arbor, MI, USA.

The orphan nuclear receptor NR2E3 is a direct transcriptional target of NRL, the key basic motif leucine zipper transcription factor that dictates rod versus cone photoreceptor cell fate in the mammalian retina. The lack of NR2E3 function in humans and in retinal degeneration rd7 mutant mouse leads to increased S-cones accompanied by rod degeneration, whereas ectopic expression of Nr2e3 in the cone-only Nrl(-/-) retina generates rod-like cells that do not exhibit any visual function. Using GFP to tag the newborn rods and by 5-bromo-2'-deoxyuridine birthdating, we demonstrate that early-born post-mitotic photoreceptor precursors in the rd7 retina express cone-specific genes. Transgenic mouse studies in the rd7 background show that Nr2e3 when expressed under the control of Crx promoter can restore rod photoreceptor function and suppress cone gene expression. Furthermore, Nr2e3 expression in photoreceptor precursors committed to be rods (driven by the Nrl promoter) could completely rescue the retinal phenotype of the rd7 mice. We conclude that excess of S-cones in the rd7 retina originate from photoreceptor precursors with a 'default' fate and not from proliferation of cones and that Nr2e3 is required to suppress the expression of S-cone genes during normal rod differentiation. These studies further support the 'transcriptional dominance' model of photoreceptor cell fate determination and provide insights into the pathogenesis of retinal disease phenotypes caused by NR2E3 mutations.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/hmg/ddr334DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3188990PMC
November 2011

A regulatory domain is required for Foxn4 activity during retinogenesis.

J Mol Neurosci 2012 Feb 24;46(2):315-23. Epub 2011 Jun 24.

Centre de Recherche des Cordeliers, INSERM UMR S872, 15, Rue de L'Ecole de Médecine, 75006, Paris, France.

Foxn4, a member of the N-family forkhead transcription factors, controls fate decision in mouse retina and spinal cord as well as in zebrafish heart. Analysis of Foxn4 amino acid sequence revealed the presence of a region homologous to the activation domain of its close relative Foxn1 in between C-terminal amino acids 402 and 455 of Foxn4 protein. The requirement of Foxn4 putative activation domain remains to be elucidated. Using a gain-of function approach in rat and chick retinal explants, we report that deletion of Foxn4 putative activation domain results in a complete loss of its activity during retinogenesis. Target promoter transcription assay indicates that this domain is critical for Foxn4 transcriptional regulatory properties in vitro. Accordingly, in chick retinal explants, this domain is required for proper regulation of target retinogenic factors expression by Foxn4. Thus, our study demonstrates that the domain between amino acids 402 and 455 is necessary for Foxn4 transcriptional activity both in vitro and in the retina.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s12031-011-9585-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3518883PMC
February 2012

Sumoylation of bZIP transcription factor NRL modulates target gene expression during photoreceptor differentiation.

J Biol Chem 2010 Aug 15;285(33):25637-44. Epub 2010 Jun 15.

Neurobiology-Neurodegeneration and Repair Laboratory, NEI, National Institutes of Health, Bethesda, Maryland 20892, USA.

Development of rod photoreceptors in the mammalian retina is critically dependent on the basic motif-leucine zipper transcription factor NRL (neural retina leucine zipper). In the absence of NRL, photoreceptor precursors in mouse retina produce only cones that primarily express S-opsin. Conversely, ectopic expression of NRL in post-mitotic precursors leads to a rod-only retina. To explore the role of signaling molecules in modulating NRL function, we identified putative sites of post-translational modification in the NRL protein by in silico analysis. Here, we demonstrate the sumoylation of NRL in vivo and in vitro, with two small ubiquitin-like modifier (SUMO) molecules attached to the Lys-20 residue. NRL-K20R and NRL-K20R/K24R sumoylation mutants show reduced transcriptional activation of Nr2e3 and rhodopsin promoters (two direct targets of NRL) in reporter assays when compared with wild-type NRL. Consistent with this, in vivo electroporation of the NRL-K20R/K24R mutant into newborn Nrl(-/-) mouse retina leads to reduced Nr2e3 activation and only a partial rescue of the Nrl(-/-) phenotype in contrast to the wild-type NRL that is able to convert cones to rod photoreceptors. Although PIAS3 (protein inhibitor of activated STAT3), an E3-SUMO ligase implicated in photoreceptor differentiation, can be immunoprecipitated with NRL, there appears to be redundancy in E3 ligases, and PIAS3 does not seem to be essential for NRL sumoylation. Our studies suggest an important role of sumoylation in fine-tuning the activity of NRL and thereby incorporating yet another layer of control in gene regulatory networks involved in photoreceptor development and homeostasis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/jbc.M110.142810DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2919127PMC
August 2010
-->