Publications by authors named "Thomas Lamonerie"

23 Publications

  • Page 1 of 1

OTX2 Non-Cell Autonomous Activity Regulates Inner Retinal Function.

eNeuro 2020 Sep/Oct;7(5). Epub 2020 Sep 4.

Centre for Interdisciplinary Research in Biology (CIRB), Collège de France, CNRS, UMR 7241, INSERM U1050, Labex MemoLife, Université PSL (Paris Sciences & Lettres), 75005 Paris, France

OTX2 is a homeoprotein transcription factor expressed in photoreceptors and bipolar cells in the retina. OTX2, like many other homeoproteins, transfers between cells and exerts non-cell autonomous effects such as promoting the survival of retinal ganglion cells that do not express the protein. Here we used a genetic approach to target extracellular OTX2 in the retina by conditional expression of a secreted single-chain anti-OTX2 antibody. Compared with control mice, the expression of this antibody by parvalbumin-expressing neurons in the retina is followed by a reduction in visual acuity in 1-month-old mice with no alteration of the retinal structure or cell type number or aspect. The a-waves and b-waves measured by electroretinogram were also indistinguishable from those of control mice, suggesting no functional deficit of photoreceptors and bipolar cells. Mice expressing the OTX2-neutralizing antibody did show a significant doubling in the flicker amplitude and a reduction in oscillatory potential, consistent with a change in inner retinal function. Our results show that interfering with OTX2 non-cell autonomous activity in the postnatal retina leads to an alteration in inner retinal cell functions and causes a deficit in visual acuity.
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http://dx.doi.org/10.1523/ENEURO.0012-19.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7477954PMC
June 2021

Developmental Requirement of Homeoprotein Otx2 for Specific Habenulo-Interpeduncular Subcircuits.

J Neurosci 2019 02 28;39(6):1005-1019. Epub 2018 Dec 28.

Université Côte d'Azur, Centre National de la Recherche Scientifique, Inserm, Institut de Biologie Valrose, 06108 Nice, France, and

The habenulo-interpeduncular system (HIPS) is now recognized as a critical circuit modulating aversion, reward, and social behavior. There is evidence that dysfunction of this circuit leads to psychiatric disorders. Because psychiatric diseases may originate in developmental abnormalities, it is crucial to investigate the developmental mechanisms controlling the formation of the HIPS. Thus far, this issue has been the focus of limited studies. Here, we explored the developmental processes underlying the formation of the medial habenula (MHb) and its unique output, the interpeduncular nucleus (IPN), in mice independently of their gender. We report that the homeobox gene is essential for the proper development of both structures. We show that MHb and IPN neurons require at different developmental stages and, in both cases, deletion leads to disruption of HIPS subcircuits. Finally, we show that Otx2 neurons tend to be preferentially interconnected. This study reveals that synaptically connected components of the HIPS, despite radically different developmental strategies, share high sensitivity to expression. Brain reward circuits are highly complex and still poorly understood. In particular, it is important to understand how these circuits form as many psychiatric diseases may arise from their abnormal development. This work shows that , a critical evolutionary conserved gene implicated in brain development and a predisposing factor for psychiatric diseases, is required for the formation of the habenulo-interpeduncular system (HIPS), an important component of the reward circuit. Otx2 deletion affects multiple processes such as proliferation and migration of HIPS neurons. Furthermore, neurons expressing are preferentially interconnected. Therefore, expression may represent a code that specifies the connectivity of functional subunits of the HIPS. Importantly, the conditional knock-out animals used in this study might represent a new genetic model of psychiatric diseases.
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http://dx.doi.org/10.1523/JNEUROSCI.1818-18.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6363926PMC
February 2019

Otx2 promotes granule cell precursor proliferation and Shh-dependent medulloblastoma maintenance in vivo.

Oncogenesis 2018 Aug 13;7(8):60. Epub 2018 Aug 13.

Université Côte d'Azur CNRS, Inserm, iBV, Institut de Biologie Valrose, Nice Cedex 2, France.

The developmental gene OTX2 is expressed by cerebellar granule cell precursors (GCPs), a cell population which undergoes massive expansion during the early postnatal period in response to sonic hedgehog (Shh). GCPs are thought to be at the origin of most medulloblastomas, a devastating paediatric cancer that arises in the developing cerebellum. OTX2 is overexpressed in all types of medulloblastomas, except in Shh-dependent type 2 medulloblastomas, although it has GCPs as cell-of-origin. This has led to the current view that OTX2 is not involved in tumorigenesis of this subgroup. How OTX2 might contribute to normal or tumoral GCP development in vivo remains unresolved. Here, we have investigated, for the first time, the physiological function of this factor in regulating proliferation and tumorigenesis in the developing mouse cerebellum. We first characterized Otx2-expressing cells in the early postnatal cerebellum and showed that they represent a unique subpopulation of highly proliferative GCPs. We next performed in vivo loss-of-function analysis to dissect out the role of Otx2 in these cells and identified a novel, Shh-independent, function for this factor in controlling postnatal GCP proliferation and cerebellum morphogenesis. Finally, we addressed the function of Otx2 in the context of type 2 medulloblastomas by directing Shh-dependent tumour formation in Otx2+ cells of the developing cerebellum and assessing the effects of Otx2 ablation in this context. We unravel an unexpected, mandatory function for Otx2 in sustaining cell proliferation and long-term maintenance of these tumours in vivo, therefore bringing unpredicted insight into the mechanisms of type 2 medulloblastoma subsistence. Together, these data pinpoint, for the first time, a crucial Shh-independent role for Otx2 in the control of proliferation of normal and tumoral granule cell precursors in vivo and make it an attractive candidate for targeted therapy in Shh-dependent medulloblastomas.
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http://dx.doi.org/10.1038/s41389-018-0070-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6087714PMC
August 2018

The caudo-ventral pallium is a novel pallial domain expressing Gdf10 and generating Ebf3-positive neurons of the medial amygdala.

Brain Struct Funct 2018 Sep 4;223(7):3279-3295. Epub 2018 Jun 4.

Université Côte d'Azur (UCA), CNRS, Inserm, Institut de Biologie Valrose (iBV), 06108, Nice, France.

In rodents, the medial nucleus of the amygdala receives direct inputs from the accessory olfactory bulbs and is mainly implicated in pheromone-mediated reproductive and defensive behaviors. The principal neurons of the medial amygdala are GABAergic neurons generated principally in the caudo-ventral medial ganglionic eminence and preoptic area. Beside GABAergic neurons, the medial amygdala also contains glutamatergic Otp-expressing neurons cells generated in the lateral hypothalamic neuroepithelium and a non-well characterized Pax6-positive population. In the present work, we describe a novel glutamatergic Ebf3-expressing neuronal subpopulation distributed within the periphery of the postero-ventral medial amygdala. These neurons are generated in a pallial domain characterized by high expression of Gdf10. This territory is topologically the most caudal tier of the ventral pallium and accordingly, we named it Caudo-Ventral Pallium (CVP). In the absence of Pax6, the CVP is disrupted and Ebf3-expressing neurons fail to be generated. Overall, this work proposes a novel model of the neuronal composition of the medial amygdala and unravels for the first time a new novel pallial subpopulation originating from the CVP and expressing the transcription factor Ebf3.
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http://dx.doi.org/10.1007/s00429-018-1687-0DOI Listing
September 2018

A new genetically engineered mouse model of choroid plexus carcinoma.

Biochem Biophys Res Commun 2018 02 12;496(2):568-574. Epub 2018 Jan 12.

Université Côte d'Azur, CNRS, Inserm, iBV, Nice, France. Electronic address:

Choroid plexus carcinomas (CPCs) are highly malignant brain tumours predominantly found in children and associated to poor prognosis. Improved therapy for these cancers would benefit from the generation of animal models. Here we have created a novel mouse CPC model by expressing a stabilised form of c-Myc (MycT58A) and inactivating Trp53 in the choroid plexus of newborn mice. This induced aberrant proliferation of choroid plexus epithelial cells, leading to aggressive tumour development and death within 150 days. Choroid plexus tumours occurred with a complete penetrance in all brain ventricles, with prevalence in the lateral and fourth ventricles. Histological and cellular analysis indicated that these tumours were CPCs resembling their human counterparts. Comparison of gene expression profiles of CPCs and non-neoplastic tissues revealed profound alterations in cell cycle regulation and DNA damage responses, suggesting that dysregulation of cell division and DNA checkpoint pathways may represent key vulnerabilities. This novel animal model of CPC provides an invaluable tool to elucidate the mechanism of CPC formation and to develop successful therapies against this devastating paediatric cancer.
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http://dx.doi.org/10.1016/j.bbrc.2017.11.192DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5880203PMC
February 2018

Mitochondrial Protection by Exogenous Otx2 in Mouse Retinal Neurons.

Cell Rep 2015 Nov 22;13(5):990-1002. Epub 2015 Oct 22.

Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon 305-701, South Korea. Electronic address:

OTX2 (orthodenticle homeobox 2) haplodeficiency causes diverse defects in mammalian visual systems ranging from retinal dysfunction to anophthalmia. We find that the retinal dystrophy of Otx2(+/GFP) heterozygous knockin mice is mainly due to the loss of bipolar cells and consequent deficits in retinal activity. Among bipolar cell types, OFF-cone bipolar subsets, which lack autonomous Otx2 gene expression but receive Otx2 proteins from photoreceptors, degenerate most rapidly in Otx2(+/GFP) mouse retinas, suggesting a neuroprotective effect of the imported Otx2 protein. In support of this hypothesis, retinal dystrophy in Otx2(+/GFP) mice is prevented by intraocular injection of Otx2 protein, which localizes to the mitochondria of bipolar cells and facilitates ATP synthesis as a part of mitochondrial ATP synthase complex. Taken together, our findings demonstrate a mitochondrial function for Otx2 and suggest a potential therapeutic application of OTX2 protein delivery in human retinal dystrophy.
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http://dx.doi.org/10.1016/j.celrep.2015.09.075DOI Listing
November 2015

Comprehensive interactome of Otx2 in the adult mouse neural retina.

Genesis 2015 Nov 23;53(11):685-94. Epub 2015 Oct 23.

Institut De Biologie Valrose, University of Nice Sophia Antipolis, UFR Sciences, CNRS, UMR7277, Inserm, U1091, Nice, F-06108, France.

The Otx2 homeodomain transcription factor exerts multiple functions in specific developmental contexts, probably through the regulation of different sets of genes. Protein partners of Otx2 have been shown to modulate its activity. Therefore, the Otx2 interactome may play a key role in selecting a precise target-gene repertoire, hence determining its function in a specific tissue. To address the nature of Otx2 interactome, we generated a new recombinant Otx2(CTAP-tag) mouse line, designed for protein complexes purification. We validated this mouse line by establishing the Otx2 interactome in the adult neural retina. In this tissue, Otx2 is thought to have overlapping function with its paralog Crx. Our analysis revealed that, in contrary to Crx, Otx2 did not develop interactions with proteins that are known to regulate phototransduction genes but showed specific partnership with factors associated with retinal development. The relationship between Otx2 and Crx in the neural retina should therefore be considered as complementarity rather than redundancy. Furthermore, study of the Otx2 interactome revealed strong associations with RNA processing and translation machineries, suggesting unexpected roles for Otx2 in the regulation of selected target genes all along the transcription/translation pathway. The Otx2(CTAP-tag) line, therefore, appears suitable for a systematic approach to Otx2 protein-protein interactions. genesis 53:685-694, 2015. © 2015 Wiley Periodicals, Inc.
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http://dx.doi.org/10.1002/dvg.22903DOI Listing
November 2015

Otx2 is a target of N-myc and acts as a suppressor of sensory development in the mammalian cochlea.

Development 2015 Aug 9;142(16):2792-800. Epub 2015 Jul 9.

Instituto de Biología y Genética Molecular, Universidad de Valladolid y Consejo Superior de Investigaciones Científicas, C/Sanz y Forés 3, Valladolid E-47003, Spain

Transcriptional regulatory networks are essential during the formation and differentiation of organs. The transcription factor N-myc is required for proper morphogenesis of the cochlea and to control correct patterning of the organ of Corti. We show here that the Otx2 gene, a mammalian ortholog of the Drosophila orthodenticle homeobox gene, is a crucial target of N-myc during inner ear development. Otx2 expression is lost in N-myc mouse mutants, and N-myc misexpression in the chick inner ear leads to ectopic expression of Otx2. Furthermore, Otx2 enhancer activity is increased by N-myc misexpression, indicating that N-myc may directly regulate Otx2. Inactivation of Otx2 in the mouse inner ear leads to ectopic expression of prosensory markers in non-sensory regions of the cochlear duct. Upon further differentiation, these domains give rise to an ectopic organ of Corti, together with the re-specification of non-sensory areas into sensory epithelia, and the loss of Reissner's membrane. Therefore, the Otx2-positive domain of the cochlear duct shows a striking competence to develop into a mirror-image copy of the organ of Corti. Taken together, these data show that Otx2 acts downstream of N-myc and is essential for patterning and spatial restriction of the sensory domain of the mammalian cochlea.
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http://dx.doi.org/10.1242/dev.122465DOI Listing
August 2015

Deletion of OTX2 in neural ectoderm delays anterior pituitary development.

Hum Mol Genet 2015 Feb 14;24(4):939-53. Epub 2014 Oct 14.

Department of Human Genetics, University of Michigan, Ann Arbor, MI 48109-5618, USA and

OTX2 is a homeodomain transcription factor that is necessary for normal head development in mouse and man. Heterozygosity for loss-of-function alleles causes an incompletely penetrant, haploinsufficiency disorder. Affected individuals exhibit a spectrum of features that range from developmental defects in eye and/or pituitary development to acephaly. To investigate the mechanism underlying the pituitary defects, we used different cre lines to inactivate Otx2 in early head development and in the prospective anterior and posterior lobes. Mice homozygous for Otx2 deficiency in early head development and pituitary oral ectoderm exhibit craniofacial defects and pituitary gland dysmorphology, but normal pituitary cell specification. The morphological defects mimic those observed in humans and mice with OTX2 heterozygous mutations. Mice homozygous for Otx2 deficiency in the pituitary neural ectoderm exhibited altered patterning of gene expression and ablation of FGF signaling. The posterior pituitary lobe and stalk, which normally arise from neural ectoderm, were extremely hypoplastic. Otx2 expression was intact in Rathke's pouch, the precursor to the anterior lobe, but the anterior lobe was hypoplastic. The lack of FGF signaling from the neural ectoderm was sufficient to impair anterior lobe growth, but not the differentiation of hormone-producing cells. This study demonstrates that Otx2 expression in the neural ectoderm is important intrinsically for the development of the posterior lobe and pituitary stalk, and it has significant extrinsic effects on anterior pituitary growth. Otx2 expression early in head development is important for establishing normal craniofacial features including development of the brain, eyes and pituitary gland.
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http://dx.doi.org/10.1093/hmg/ddu506DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4834879PMC
February 2015

Head formation: OTX2 regulates Dkk1 and Lhx1 activity in the anterior mesendoderm.

Development 2014 Oct 17;141(20):3859-67. Epub 2014 Sep 17.

Embryology Unit, Children's Medical Research Institute, Locked Bag 23, Wentworthville, New South Wales 2145, Australia Discipline of Medicine, Sydney Medical School, University of Sydney, New South Wales 2006, Australia.

The Otx2 gene encodes a paired-type homeobox transcription factor that is essential for the induction and the patterning of the anterior structures in the mouse embryo. Otx2 knockout embryos fail to form a head. Whereas previous studies have shown that Otx2 is required in the anterior visceral endoderm and the anterior neuroectoderm for head formation, its role in the anterior mesendoderm (AME) has not been assessed specifically. Here, we show that tissue-specific ablation of Otx2 in the AME phenocopies the truncation of the embryonic head of the Otx2 null mutant. Expression of Dkk1 and Lhx1, two genes that are also essential for head formation, is disrupted in the AME of the conditional Otx2-deficient embryos. Consistent with the fact that Dkk1 is a direct target of OTX2, we showed that OTX2 can interact with the H1 regulatory region of Dkk1 to activate its expression. Cross-species comparative analysis, RT-qPCR, ChIP-qPCR and luciferase assays have revealed two conserved regions in the Lhx1 locus to which OTX2 can bind to activate Lhx1 expression. Abnormal development of the embryonic head in Otx2;Lhx1 and Otx2;Dkk1 compound mutant embryos highlights the functional intersection of Otx2, Dkk1 and Lhx1 in the AME for head formation.
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http://dx.doi.org/10.1242/dev.114900DOI Listing
October 2014

Otx2 ChIP-seq reveals unique and redundant functions in the mature mouse retina.

PLoS One 2014 18;9(2):e89110. Epub 2014 Feb 18.

Institut de Biologie Valrose, University of Nice Sophia Antipolis, CNRS UMR7277, Inserm U1091, Nice, France.

During mouse retinal development and into adulthood, the transcription factor Otx2 is expressed in pigment epithelium, photoreceptors and bipolar cells. In the mature retina, Otx2 ablation causes photoreceptor degeneration through a non-cell-autonomous mechanism involving Otx2 function in the supporting RPE. Surprisingly, photoreceptor survival does not require Otx2 expression in the neural retina, where the related Crx homeobox gene, a major regulator of photoreceptor development, is also expressed. To get a deeper view of mouse Otx2 activities in the neural retina, we performed chromatin-immunoprecipitation followed by massively parallel sequencing (ChIP-seq) on Otx2. Using two independent ChIP-seq assays, we identified consistent sets of Otx2-bound cis-regulatory elements. Comparison with our previous RPE-specific Otx2 ChIP-seq data shows that Otx2 occupies different functional domains of the genome in RPE cells and in neural retina cells and regulates mostly different sets of genes. To assess the potential redundancy of Otx2 and Crx, we compared our data with Crx ChIP-seq data. While Crx genome occupancy markedly differs from Otx2 genome occupancy in the RPE, it largely overlaps that of Otx2 in the neural retina. Thus, in accordance with its essential role in the RPE and its non-essential role in the neural retina, Otx2 regulates different gene sets in the RPE and the neural retina, and shares an important part of its repertoire with Crx in the neural retina. Overall, this study provides a better understanding of gene-regulatory networks controlling photoreceptor homeostasis and disease.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0089110PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3928427PMC
October 2014

Purkinje cells and Bergmann glia are primary targets of the TRα1 thyroid hormone receptor during mouse cerebellum postnatal development.

Development 2014 Jan;141(1):166-75

Université de Lyon, CNRS, INRA, Université Claude Bernard Lyon 1, École Normale Supérieure de Lyon, Institut de Génomique Fonctionnelle de Lyon, F-69364 Lyon, Cedex 07, France.

Thyroid hormone is necessary for normal development of the central nervous system, as shown by the severe mental retardation syndrome affecting hypothyroid patients with low levels of active thyroid hormone. The postnatal defects observed in hypothyroid mouse cerebellum are recapitulated in mice heterozygous for a dominant-negative mutation of Thra, the gene encoding the ubiquitous TRα1 receptor. Using CRE/loxP-mediated conditional expression approach, we found that this mutation primarily alters the differentiation of Purkinje cells and Bergmann glia, two cerebellum-specific cell types. These primary defects indirectly affect cerebellum development in a global manner. Notably, the inward migration and terminal differentiation of granule cell precursors is impaired. Therefore, despite the broad distribution of its receptors, thyroid hormone targets few cell types that exert a predominant role in the network of cellular interactions that govern normal cerebellum maturation.
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http://dx.doi.org/10.1242/dev.103226DOI Listing
January 2014

Loss of Otx2 in the adult retina disrupts retinal pigment epithelium function, causing photoreceptor degeneration.

J Neurosci 2013 Jun;33(24):9890-904

Institut de Biologie Valrose, University of Nice Sophia Antipolis, UFR Sciences, Nice F-06108, France.

Photoreceptors are specialized neurons of the retina that receive nursing from the adjacent retinal pigment epithelium (RPE). Frequent in the elderly, photoreceptor loss can originate from primary dysfunction of either cell type. Despite intense interest in the etiology of these diseases, early molecular actors of late-onset photoreceptor degeneration remain elusive, mostly because of the lack of dedicated models. Conditional Otx2 ablation in the adult mouse retina elicits photoreceptor degeneration, providing a new model of late-onset neuronal disease. Here, we use this model to identify the earliest events after Otx2 ablation. Electroretinography and gene expression analyses suggest a nonautonomous, RPE-dependent origin for photoreceptor degeneration. This is confirmed by RPE-specific ablation of Otx2, which results in similar photoreceptor degeneration. In contrast, constitutive Otx2 expression in RPE cells prevents degeneration of photoreceptors in Otx2-ablated retinas. We use chromatin immunoprecipitation followed by massive sequencing (ChIP-seq) analysis to identify the molecular network controlled in vivo by Otx2 in RPE cells. We uncover four RPE-specific functions coordinated by Otx2 that underpin the cognate photoreceptor degeneration. Many direct Otx2 target genes are associated with human retinopathies, emphasizing the significance of the model. Importantly, we report a secondary genetic response after Otx2 ablation, which largely precedes apoptosis of photoreceptors, involving inflammation and stress genes. These findings thus provide novel general markers for clinical detection and prevention of neuronal cell death.
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http://dx.doi.org/10.1523/JNEUROSCI.1099-13.2013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6618395PMC
June 2013

The homeobox gene Otx2 in development and disease.

Exp Eye Res 2013 Jun 21;111:9-16. Epub 2013 Mar 21.

Department of Ophthalmology, Hôpital Universitaire Des Enfants Reine Fabiola (HUDERF), Université Libre de Bruxelles (ULB), Avenue Jean Joseph Crocq 15, 1020 Brussels, Belgium.

The Otx2 gene encodes a transcription factor essential for the normal development of brain, cerebellum, pineal gland, and eye. In the retina, Otx2 has essential functions from early embryogenesis to adulthood. As soon as the optic vesicle is formed, the gene is required for retinal pigment epithelium specification. Otx2 is also a key regulator of photoreceptor genesis and differentiation, and is required after birth for bipolar cells terminal maturation. Otx2 expression is maintained in the differentiated retina wherein the gene is critical for the outer retina maintenance. In the visual cortex, the gene modulates the neuronal plasticity through a paracrine mechanism. OTX2 heterozygous mutations in humans have been linked to severe ocular malformations associated with brain abnormalities and pituitary dysfunction. Recent studies have also established the OTX2 gene as an oncogene for medulloblastoma, a malignant brain tumour originating in the cerebellum.
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http://dx.doi.org/10.1016/j.exer.2013.03.007DOI Listing
June 2013

ES cell-derived renewable and functional midbrain dopaminergic progenitors.

Proc Natl Acad Sci U S A 2011 Jun 23;108(23):9703-8. Epub 2011 May 23.

Molecular Neurobiology Laboratory and Department of Psychiatry and Program in Neuroscience, Harvard Stem Cell Institute, McLean Hospital/Harvard Medical School, Belmont, MA 02478, USA.

During early development, midbrain dopaminergic (mDA) neuronal progenitors (NPs) arise from the ventral mesencephalic area by the combined actions of secreted factors and their downstream transcription factors. These mDA NPs proliferate, migrate to their final destinations, and develop into mature mDA neurons in the substantia nigra and the ventral tegmental area. Here, we show that such authentic mDA NPs can be efficiently isolated from differentiated ES cells (ESCs) using a FACS method combining two markers, Otx2 and Corin. Purified Otx2(+)Corin(+) cells coexpressed other mDA NP markers, including FoxA2, Lmx1b, and Glast. Using optimized culture conditions, these mDA NPs continuously proliferated up to 4 wk with almost 1,000-fold expansion without significant changes in their phenotype. Furthermore, upon differentiation, Otx2(+)Corin(+) cells efficiently generated mDA neurons, as evidenced by coexpression of mDA neuronal markers (e.g., TH, Pitx3, Nurr1, and Lmx1b) and physiological functions (e.g., efficient DA secretion and uptake). Notably, these mDA NPs differentiated into a relatively homogenous DA population with few serotonergic neurons. When transplanted into PD model animals, aphakia mice, and 6-OHDA-lesioned rats, mDA NPs differentiated into mDA neurons in vivo and generated well-integrated DA grafts, resulting in significant improvement in motor dysfunctions without tumor formation. Furthermore, grafted Otx2(+)Corin(+) cells exhibited significant migratory function in the host striatum, reaching >3.3 mm length in the entire striatum. We propose that functional and expandable mDA NPs can be efficiently isolated by this unique strategy and will serve as useful tools in regenerative medicine, bioassay, and drug screening.
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http://dx.doi.org/10.1073/pnas.1016443108DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3111266PMC
June 2011

Otx2 gene deletion in adult mouse retina induces rapid RPE dystrophy and slow photoreceptor degeneration.

PLoS One 2010 Jul 21;5(7):e11673. Epub 2010 Jul 21.

Institut de Génomique Fonctionnelle de Lyon, Centre National de la Recherche Scientifique, Ecole Normale Supérieure de Lyon, Université de Lyon, Lyon, France.

Background: Many developmental genes are still active in specific tissues after development is completed. This is the case for the homeobox gene Otx2, an essential actor of forebrain and head development. In adult mouse, Otx2 is strongly expressed in the retina. Mutations of this gene in humans have been linked to severe ocular malformation and retinal diseases. It is, therefore, important to explore its post-developmental functions. In the mature retina, Otx2 is expressed in three cell types: bipolar and photoreceptor cells that belong to the neural retina and retinal pigment epithelium (RPE), a neighbour structure that forms a tightly interdependent functional unit together with photoreceptor cells.

Methodology/principal Findings: Conditional self-knockout was used to address the late functions of Otx2 gene in adult mice. This strategy is based on the combination of a knock-in CreERT2 allele and a floxed allele at the Otx2 locus. Time-controlled injection of tamoxifen activates the recombinase only in Otx2 expressing cells, resulting in selective ablation of the gene in its entire domain of expression. In the adult retina, loss of Otx2 protein causes slow degeneration of photoreceptor cells. By contrast, dramatic changes of RPE activity rapidly occur, which may represent a primary cause of photoreceptor disease.

Conclusions: Our novel mouse model uncovers new Otx2 functions in adult retina. We show that this transcription factor is necessary for long-term maintenance of photoreceptors, likely through the control of specific activities of the RPE.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0011673PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2908139PMC
July 2010

FGFRL1 is a neglected putative actor of the FGF signalling pathway present in all major metazoan phyla.

BMC Evol Biol 2009 Sep 9;9:226. Epub 2009 Sep 9.

Departament de Genètica, Facultat de Biologia, Universitat de Barcelona, Av. Diagonal 645, edifici annex, 1a planta, 08028 Barcelona, España.

Background: Fibroblast Growth Factors (FGF) and their receptors are well known for having major implications in cell signalling controlling embryonic development. Recently, a gene coding for a protein closely related to FGFRs (Fibroblast Growth Factor Receptors) called FGFR5 or FGFR-like 1 (FGFRL1), has been described in vertebrates. An orthologous gene was also found in the cephalochordate amphioxus, but no orthologous genes were found by the authors in other non-vertebrate species, even if a FGFRL1 gene was identified in the sea urchin genome, as well as a closely related gene, named nou-darake, in the planarian Dugesia japonica. These intriguing data of a deuterostome-specific gene that might be implicated in FGF signalling prompted us to search for putative FGFRL1 orthologues in the completely sequenced genomes of metazoans.

Results: We found FGFRL1 genes in the cnidarian Nematostella vectensis as well as in many bilaterian species. Our analysis also shows that FGFRL1 orthologous genes are linked in the genome with other members of the FGF signalling pathway from cnidarians to bilaterians (distance < 10 Mb). To better understand the implication of FGFRL1 genes in chordate embryonic development, we have analyzed expression patterns of the amphioxus and the mouse genes by whole mount in situ hybridization. We show that some homologous expression territories can be defined, and we propose that FGFRL1 and FGF8/17/18 were already co-expressed in the pharyngeal endoderm in the ancestor of chordates.

Conclusion: Our work sheds light on the existence of a putative FGF signalling pathway actor present in the ancestor of probably all metazoans, the function of which has received little attention until now.
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http://dx.doi.org/10.1186/1471-2148-9-226DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2754479PMC
September 2009

A new GFP-tagged line reveals unexpected Otx2 protein localization in retinal photoreceptors.

BMC Dev Biol 2007 Nov 2;7:122. Epub 2007 Nov 2.

IGFL, UMR CNRS 5242-INRA 1237-ENS, IFR128 Lyon-Gerland, 46 allée d'Italie, 69364 Lyon Cedex 07, France.

Background: Dynamic monitoring of protein expression and localization is fundamental to the understanding of biological processes. The paired-class homeodomain-containing transcription factor Otx2 is essential for normal head and brain development in vertebrates. Recent conditional knockout studies have pointed to multiple roles of this protein during late development and post-natal life. Yet, later expression and functions remain poorly characterized as specific reagents to detect the protein at any stage of development are still missing.

Results: We generated a new mouse line harbouring an insertion of the GFP gene within the Otx2 coding sequence to monitor the gene activity while preserving most of its functions. Our results demonstrate that this line represents a convenient tool to capture the dynamics of Otx2 gene expression from early embryonic stages to adulthood. In addition, we could visualize the intracellular location of Otx2 protein. In the retina, we reinterpret the former view of protein distribution and show a further level of regulation of intranuclear protein localization, which depends on the cell type.

Conclusion: The GFP-tagged Otx2 mouse line fully recapitulates previously known expression patterns and brings additional accuracy and easiness of detection of Otx2 gene activity. This opens up the way to live imaging of a highly dynamic actor of brain development and can be adapted to any mutant background to probe for genetic interaction between Otx2 and the mutated gene.
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http://dx.doi.org/10.1186/1471-213X-7-122DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2204009PMC
November 2007

OTX5 regulates pineal expression of the zebrafish REV-ERB alpha through a new DNA binding site.

Mol Endocrinol 2008 Jan 13;22(1):23-32. Epub 2007 Sep 13.

Université de Lyon, Institut de Génomique Fonctionnelle de Lyon, Unité Mixte de Recherche 5242 du Centre National de la Recherche Scientifique, 69364 Lyon Cedex 07, France.

The pineal gland plays a central role in the photoneuroendocrine system and acts as a photosensory organ in lower vertebrates. The orphan nuclear receptor Rev-erbalpha (NR1D1) has previously been shown to be expressed in the pineal and to be regulated with a robust circadian rhythm during zebrafish embryogenesis. This early pineal expression is under the control of the transcription factor Orthodenticle homeobox 5 (Otx5). In this paper, we show that Otx5 regulates the second zfRev-erbalpha promoter, ZfP2. Despite the absence of a classical Otx-binding site within ZfP2, this regulation depends on the integrity of the Otx5 homeodomain. Mapping experiments as well as EMSAs show that this interaction between Otx5 and ZfP2 depends on a noncanonical bipartite Otx-binding site (GANNCTTA and TAAA) that we called pineal expression related element (PERE). We showed that PERE is necessary for pineal expression in vivo by injecting zebrafish embryos with wild type and mutated versions of zfRev-erbalpha promoter fused to green fluorescent protein. Interestingly, PERE is found upstream of other genes expressed in the pineal gland, suggesting that it may play an important role in governing pineal expression. Our data establish that PERE is a novel cis-acting element contributing to pineal-specific gene expression and to Otx target gene regulation.
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http://dx.doi.org/10.1210/me.2007-0170DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5419633PMC
January 2008

Temporal and spatial delineation of mouse Otx2 functions by conditional self-knockout.

EMBO Rep 2006 Aug 14;7(8):824-30. Epub 2006 Jul 14.

BMC, UMR CNRS 5161-INRA 1237-ENS, IFR128 Lyon-Gerland, 46 allée d'Italie, 69364 Lyon Cedex 07, France.

To identify the independent spatial and temporal activities of the essential developmental gene the Otx2, the germline mutation of which is lethal at embryonic day 8.5, we floxed one allele and substituted the other with an inducible CreER recombinase gene. This makes 'trans' self-knockout possible at any developmental stage. The transient action of tamoxifen pulses allows time-course mutation. We demonstrate efficient temporal knockout and demarcate spatio-temporal windows in which Otx2 controls the head, brain structures and body development.
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http://dx.doi.org/10.1038/sj.embor.7400751DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1525150PMC
August 2006

Molecular dissection reveals decreased activity and not dominant negative effect in human OTX2 mutants.

J Mol Med (Berl) 2006 Jul 11;84(7):604-15. Epub 2006 Apr 11.

LBMC, ENS-Lyon, IFR128 Lyon-Gerland, 46 allée d'Italie, 69364, Lyon, Cedex 07, France.

The paired-type homeodomain transcription factor Otx2 is essential for forebrain and eye development. Severe ocular malformations in humans have recently been associated with heterozygous OTX2 mutations. To document the molecular defects in human mutants, Otx2 structural characterization was carried out. A collection of deletion and point mutants was created to perform transactivation, DNA binding, and subcellular localization analyses. Transactivation was ascribed to both N- and C-termini of the protein, and DNA binding to the minimal homeodomain, where critical amino acid residues were identified. Acute nuclear localization appeared controlled by a nuclear localization sequence located within the homeodomain which acts in conjunction with a novel nuclear retention domain that we unraveled located in the central part of the protein. This region, which is poorly conserved among Otx proteins, was also endowed with dominant negative activity suggesting that it might confer unique properties to Otx2. Molecular diagnostic of human mutant OTX2 proteins discriminates hypomorphic and loss of function mutations from other mutations that may not be relevant to ocular pathology.
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http://dx.doi.org/10.1007/s00109-006-0048-2DOI Listing
July 2006

Alternative usage of Otx2 promoters during mouse development.

Dev Dyn 2005 May;233(1):154-60

LBMC, ENS-Lyon, IFR128 Lyon-Gerland, 69364 Lyon Cedex 07, France.

Our previous structural analysis of mouse Otx2 transcripts has revealed the existence of three different promoters and suggested that the corresponding mRNAs could exhibit specific expression patterns. Here, we analyze the precise dynamics of their expression throughout mouse development. Their spatial distribution was determined by isoform-specific in situ hybridization and their relative abundance by real-time reverse transcriptase-polymerase chain reaction. Although the three promoters may be used in the same areas, we show that transcription preferentially occurs from the proximal promoter at onset of gene activity in early embryogenesis, and switches to the more distal one in most of the sites of expression in the adult brain. During gestation, their relative utilization becomes inverted. The third promoter, which shows no activity in embryonic stem cells and is moderately expressed during embryogenesis, is mostly used in specific areas derived from the rostral part of the neural tube.
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http://dx.doi.org/10.1002/dvdy.20287DOI Listing
May 2005

New Otx2 mRNA isoforms expressed in the mouse brain.

J Neurochem 2003 Feb;84(4):840-53

LBMC, ENS-Lyon, 46 allée d'Italie, 69364 Lyon cedex 07, France.

The mouse Otx2 gene is essential throughout head and brain development, from anterior-posterior polarity determination and neuroectoderm induction to post-natal sensory organ maturation. These numerous activities must rely on a very finely tuned regulation of expression. In order to understand the molecular control of the Otx2 gene, we set out to isolate its promoter. During this quest, we identified three remote transcription start sites, two defining two new upstream exons and one mapping within the previously reported first exon. The three transcripts differed in their 5' non-coding region but encoded the same protein. The transcription start nucleotides of each mRNA species have been mapped by RNase protection assays and by an RNA circularization technique. We have demonstrated that they are all used and linked to functional promoters. In addition to leader versatility, we also detected alternative splicing within the coding sequence that gives rise to a new protein endowed with an 8 amino-acid insertion upstream of the homeodomain. Combined analysis of the relative abundance of Otx2 mRNA isoforms in representative tissues and in situ hybridization studies revealed distinct spatial and temporal, although partially overlapping, expression patterns of the mRNA isoforms. These findings provide new clues to a better understanding of the relationships between Otx2 gene architecture and its complex regulatory requirements.
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http://dx.doi.org/10.1046/j.1471-4159.2003.01583.xDOI Listing
February 2003
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