Genomic Analysis Reveals Disruption of Striatal Neuronal Development and Therapeutic Targets in Human Huntington's Disease Neural Stem Cells.

We utilized induced pluripotent stem cells (iPSCs) derived from Huntington's disease (HD) patients as a human model of HD and determined that the disease phenotypes only manifest in the differentiated neural stem cell (NSC) stage, not in iPSCs. To understand the molecular basis for the CAG repeat expansion-dependent disease phenotypes in NSCs, we performed transcriptomic analysis of HD iPSCs and HD NSCs compared to isogenic controls. Differential gene expression and pathway analysis pointed to transforming growth factor β (TGF-β) and netrin-1 as the top dysregulated pathways. Using data-driven gene coexpression network analysis, we identified seven distinct coexpression modules and focused on two that were correlated with changes in gene expression due to the CAG expansion. Our HD NSC model revealed the dysregulation of genes involved in neuronal development and the formation of the dorsal striatum. The striatal and neuronal networks disrupted could be modulated to correct HD phenotypes and provide therapeutic targets

A Comprehensive Model and Modulation of Cellular Signaling Involved in Early Mammary Development and Aggressive Cancer Using a Novel Recombinant Protein of the G3 Domain of Laminin-5

The mammary gland is a unique and specialized epidermal organ; mammary organogenesis begins in the embryo but is not fully complete until puberty. As such, formation of the mammary gland depends on temporally and spatially regulated developmental steps that require coordination of multiple biological and cell signaling processes; many of which have parallels with cancer development. Research describing the events that occur between birth and puberty is lacking and little is known about human breast development of youth. Since mammary gland development requires a coordinated balance between cell growth, proliferation, and apoptosis, it is critical to understand which signaling pathways are utilized to relay developmental signals, and how these pathways and their targets interact and cooperate with age. Additionally, interactions between integrin molecules and their laminin ligands, especially Laminin-5 (Ln-5; also known as Laminin-332), regulate multiple facets of both embryonic development and tumor growth, invasion, and metastasis. α6β4 integrin serves as a marker to detect distant metastases in the early stages of specific malignancies and β4 integrin overexpression has been found in basal-like breast cancers, correlating with aggressiveness to institute a prognostic β4 signature that increases with tumor grade. The mechanism α6β4 integrin utilizes to modulate oncogenic signaling through association with Ln-5 molecules in the ECM is the basis for the recombinant protein (rG3, the third of five G domains of Ln-5) produced for the work reported in this dissertation. Here, it is shown there are specific transcriptional differences and a unique interaction of a gene set over time that contributes to postnatal mammary gland development, and this model clearly shares similarities and signaling pathways with oncogenic development. Especially important are pathways of the adaptive and innate immunities, ECM remodeling and integrin interactions, and extrinsic and intrinsic TP53-mediated apoptosis, greater understanding of which could lead to early detection of potential tumorigenic growth and identification of potential treatment avenues. Presented is a comprehensive model of early mammary development along with several panels of biomarkers that possess a role in normal mammary development, are involved in aggressive cancers, and are affected by apoptosis induced by rG3 treatment. rG3 has proven to be a valuable tool to study apoptotic pathways and the crosstalk among those pathways

A gene network regulated by FGF signalling during ear development

During development cell commitment is regulated by inductive signals that are tightly controlled in time and space. In response, cells activate specific programmes, but the transcriptional circuits that maintain cell identity in a changing signalling environment are often poorly understood. Specification of inner ear progenitors is initiated by FGF signalling. Here, we establish the genetic hierarchy downstream of FGF by systematic analysis of many ear factors combined with a network inference approach. We show that FGF rapidly activates a small circuit of transcription factors forming positive feedback loops to stabilise otic progenitor identity. Our predictive network suggests that subsequently, transcriptional repressors ensure the transition of progenitors to mature otic cells, while simultaneously repressing alternative fates. Thus, we reveal the regulatory logic that initiates ear formation and highlight the hierarchical organisation of the otic gene network

Mol Psychiatry

Autism spectrum disorders (ASD) are common, complex and heterogeneous neurodevelopmental disorders. Cellular and molecular mechanisms responsible for ASD pathogenesis have been proposed based on genetic studies, brain pathology and imaging, but a major impediment to testing ASD hypotheses is the lack of human cell models. Here, we reprogrammed fibroblasts to generate induced pluripotent stem cells, neural progenitor cells (NPCs) and neurons from ASD individuals with early brain overgrowth and non-ASD controls with normal brain size. ASD-derived NPCs display increased cell proliferation because of dysregulation of a \u3b2-catenin/BRN2 transcriptional cascade. ASD-derived neurons display abnormal neurogenesis and reduced synaptogenesis leading to functional defects in neuronal networks. Interestingly, defects in neuronal networks could be rescued by insulin growth factor 1 (IGF-1), a drug that is currently in clinical trials for ASD. This work demonstrates that selection of ASD subjects based on endophenotypes unraveled biologically relevant pathway disruption and revealed a potential cellular mechanism for the therapeutic effect of IGF-1.KL2 TR000099/NCATS NIH HHS/National Center for Advancing Translational Sciences/United StatesDP2 OD006495/ODCDC CDC HHS/Office of the Director/United StatesR01 MH113924/NIMH NIH HHS/National Institute of Mental Health/United StatesR01 MH100175/NIMH NIH HHS/National Institute of Mental Health/United StatesR01 MH094753/NIMH NIH HHS/National Institute of Mental Health/United StatesR00 MH101634/NIMH NIH HHS/National Institute of Mental Health/United StatesU19 MH107367/NIMH NIH HHS/National Institute of Mental Health/United States2017-05-23T00:00:00Z27378147PMC52159916525vault:3354

The Wnt Receptor Ryk Reduces Neuronal and Cell Survival Capacity by Repressing FOXO Activity During the Early Phases of Mutant Huntingtin Pathogenicity

The Wnt receptor Ryk is an evolutionary-conserved protein important during neuronal differentiation through several mechanisms, including γ-secretase cleavage and nuclear translocation of its intracellular domain (Ryk-ICD). Although the Wnt pathway may be neuroprotective, the role of Ryk in neurodegenerative disease remains unknown. We found that Ryk is up-regulated in neurons expressing mutant huntingtin (HTT) in several models of Huntington's disease (HD). Further investigation in Caenorhabditis elegans and mouse striatal cell models of HD provided a model in which the early-stage increase of Ryk promotes neuronal dysfunction by repressing the neuroprotective activity of the longevity-promoting factor FOXO through a noncanonical mechanism that implicates the Ryk-ICD fragment and its binding to the FOXO co-factor β-catenin. The Ryk-ICD fragment suppressed neuroprotection by lin-18/Ryk loss-of-function in expanded-polyQ nematodes, repressed FOXO transcriptional activity, and abolished β-catenin protection of mutant htt striatal cells against cell death vulnerability. Additionally, Ryk-ICD was increased in the nucleus of mutant htt cells, and reducing γ-secretase PS1 levels compensated for the cytotoxicity of full-length Ryk in these cells. These findings reveal that the Ryk-ICD pathway may impair FOXO protective activity in mutant polyglutamine neurons, suggesting that neurons are unable to efficiently maintain function and resist disease from the earliest phases of the pathogenic process in HD. © 2014 Tourette et al

Systems Genome:Coordinated Gene Activity Networks, Recurring Coordination Modules, and Genome Homeostasis in Developing Neurons

Simple Summary: A synchronized global genome is a flexible, homeostatic system that underwrites ontogenic development and deprograming in disease. Abstract: As human progenitor cells differentiate into neurons, the activities of many genes change; these changes are maintained within a narrow range, referred to as genome homeostasis. This process, which alters the synchronization of the entire expressed genome, is distorted in neurodevelopmental diseases such as schizophrenia. The coordinated gene activity networks formed by altering sets of genes comprise recurring coordination modules, governed by the entropy-controlling action of nuclear FGFR1, known to be associated with DNA topology. These modules can be modeled as energy-transferring circuits, revealing that genome homeostasis is maintained by reducing oscillations (noise) in gene activity while allowing gene activity changes to be transmitted across networks; this occurs more readily in neuronal committed cells than in neural progenitors. These findings advance a model of an “entangled” global genome acting as a flexible, coordinated homeostatic system that responds to developmental signals, is governed by nuclear FGFR1, and is reprogrammed in disease

Tinkering with cusp patterning : Developmental Genetic Mechanisms in Mouse Molar Development

Teeth display considerable morphological variability, which mammals have been able to use to their advantage. Consequently, mammal teeth provide a bountiful research subject that combines information on development, functional proper-ties, and thanks to their durable substance, evolutionary history. This thesis work is focused on the patterning of cusps, the peaks that form the shape of the tooth crown, in the mouse. Mouse tooth development has been studied extensively and offers a wide variety of established methods, including culture of embryonic teeth, which allows their observation and manipulation, and the mapping of gene expression patterns and protein distributions on histological sections. It has been established that teeth develop through a series of inductive interactions between the epithelium and the mesenchyme. The interactions are mediated by signalling molecules mostly belonging to the Wnt, Bmp, Fgf, and Shh families and are used similarly in the development of other organs. The growth of a tooth is controlled by epithelial signalling centres called enamel knots, each of which gives rise to a cusp. The patterning of enamel knots, and thus of cusps, can be modelled with reaction-diffusion dynamics, which suggests the patterning to be robust against interference yet capable of propagating change. As a semi-independent developmental module, teeth can vary without affecting the rest of the organism, an assumed prerequisite for evolvability. However, the use of tooth development in evolutionary studies has been hampered by a lack of mutations and manipulations causing small-scale variation. We have explored the dynamics of cusp patterning by studying mouse mutants with altered cusp patterns and by producing cusp pattern variation in cultured molars. In addition to taking advantage of established methods, we have shown Shhwt/GFPcre reporter molars to allow real-time observation of cusp patterning in culture, derived quantified data from developing molars, and imaged their three-dimensional structure at cellular resolution with X-ray scanning. Our results indicate that cusp patterning is controlled by feedback inhibition of enamel knot differentiation, and we identify Bmp, Activin, Eda, and Fgf20 as activators, and ectodin and Shh as inhibitors of differentiation. Each of these has slightly different functions and the correct regulation of all of these is required for normal cusp patterning. Bmp and ectodin, and Eda and Shh, seem to form feedback loops providing developmental stability. The manipulation of Eda signalling provided an opportunity to quantify development, revealing that variation increased in a linear fashion the further one deviated from the wild type level of signalling. Our results support the use of reaction-diffusion dynamics in modelling cusp patterning, but they also show that growth dynamics play an equally important role. Consequently, the evolution of crown shape can be followed cusp by cusp, and the developmental order of enamel knot induction closely corresponds to the evolutionary order of cusp appearance. Thus the mechanisms of molar development can be assumed to restrict, or channel, variation available to selection. In agreement with this, most of the molar features we generated have counterparts in extinct or extant rodent species. A general trend in evolution, evident also in molars, has been an increase in complexity. In the absence of experimental ways to repeat this phenomenon, its dynamics have remained elusive. In tuning Eda, Activin, and Shh signalling we found that an increase in cusp number correlated with the number of signalling pathways tuned simultaneously. Though intuitively obvious, the result had not previously been reported. Should an increase in complexity require multiple simultaneous changes in development as a rule, the overwhelming majority of reports on decreasing complexity, typically studying the effects of a single change at a time, would be explained. In conclusion, our results provide new information on the developmental genetic mechanisms of cusp patterning, how they provide developmental stability, and what kind of evolutionary constraints they cause.Hampaiston muuntelevuus on ollut nisäkkäiden keskeinen kilpailuvaltti evoluutiossa. Erityisesti takahampaiden purupinnan nystermien koko, muoto ja kaavoitus vaihtelee suuresti lajien välillä, heijastaen kunkin lajin ruokavaliota. Kovan rakenteensa ansiosta hampaiden purupinnassa tapahtuneita muutoksia voidaan tutkia paitsi nykyisen lajikirjon, myös kattavan fossiiliaineiston avulla. Lisäksi hampaan sikiöaikaista kehitystä on tutkittu pitkään käyttäen koe-eläimenä hiirtä, jonka takahampaat ovat melko samankaltaiset ihmisen takahampaiden kanssa. Käytettävissä on siis paljon taustatietoa ja monta vakiintunutta tutkimusmenetelmää. Tämän vuoksi hampaat tarjoavat tutkimusmallin, jossa voidaan ainutlaatuisella tavalla yhdistää tietoa toiminnallisista ominaisuuksista, evoluutiohistoriasta, ja yksilönkehityksestä. Evoluution yleisimpänä mekanismina pidetään geeniverkostojen säätelyssä tapahtuvia hienovaraisia muutoksia, jotka yksilönkehityksen kautta johtavat ilmiasun muuntumiseen. Olemme tutkineet tätä ilmiötä kokeellisesti muuntogeenisillä hiirillä ja peukaloimalla kudosviljelyolosuhteissa kasvatettavien hiiren takahampaiden kehitystä. Näin olemme säädelleet hampaan kehityksessä normaalisti käytössä olevien viestintäpolkujen aktiivisuutta, nimellisesti Bmp, Ectodin, Ectodysplasin, Activin, Sonic hedgehog, ja Fibroblast growth factor 20 välitteistä viestintää. Kudosviljelyssä kasvatettujen takahampaiden kehitys kvantifioitiin mittaamalla kehitystä kuvaavat suureet päivittäin otetuista valokuvista. Solutason muutokset kohdegeenien ilmenemisessä osoitettiin in situ hybridisaatiolla, solunjakautuminen ja solukuolema puolestaan immunohistokemiallisilla värjäyksillä. Purupinnan kolmiulotteinen muoto visualisoitiin mineralisoitumattomista hampaista röntgen-skannauksella ja mineralisoituneista hampaista laser-konfokaali-skannauksella. Tuloksemme osoittavat, että Bmp, Activin, Ectodysplasin, ja Fgf20 vaikuttavat nystermien indusoitumiseen positiivisesti, kun taas ectodin ja Shh vaikuttavat negatiivisesti. Näistä Bmp ja ectodin, ja Ectodysplasin ja Shh, näyttävät muodostavan kaksi negatiivista takaisinkytkentä-silmukkaa, jotka vakauttavat hampaan kehitystä pieniä häiriöitä vastaan. Ectodysplasin-viestintää pystyttiin säätämään vähitellen, jolloin paljastui, että hampaan muuntelu lisääntyy sitä suuremmaksi, mitä kauemmaksi normaalitasosta Ectodysplasin-viestintä pakotetaan. Täten Ectodysplasin-viestinnän tason muuttuminen on saattanut olla tärkeää hampaan, tai muiden pinta-elinten, kuten kynsien ja hiusten, evoluutiohistoriassa. Lisäksi huomasimme, että ylimääräisten nystermien määrä korreloi peukaloitujen viestintäpolkujen määrän kanssa. Tuloksemme vahvistavat pitkään vallinneen teorian, että rakenteiden monimutkaisuuden lisääminen vaatii useita kehityksellisiä muutoksia, kun taas rakenteiden yksinkertaistaminen näyttäisi olevan helpompaa. Rakenteiden monimutkaisuuden lisääntyminen on ollut vallitseva suuntaus evoluutiossa, joten ilmiön toistaminen kokeellisesti avaa uusia, mielenkiintoisia tutkimusmahdollisuuksia. Yhteenvetona, tutkimuksemme ovat selventäneet miten hampaan kehityksen dynamiikka on puskuroitu tietynlaisia geneettisiä muutoksia kestäväksi, kun taas toisenlaiset muutokset tuottavat suhteettoman suuria seurauksia. Tuloksemme osoittavat, että hampaan kehityksen mekanismit ovat vinouttaneet luonnonvalinnalle tarjolla olevaa muotojen valikoimaa vaikuttaen siten evoluution kulkuun

Gain of gene regulatory network interconnectivity at the origin of vertebrates

Signaling pathways control a large number of gene regulatory networks (GRNs) during animal development, acting as major tools for body plan formation [A. Pires-daSilva, R. J. Sommer, Nat. Rev. Genet. 4, 39-49 (2003)], although only a few of these pathways operate during this period [J. J. Sanz-Ezquerro, A. E. Munsterberg, € S. Stricker, Front. Cell Dev. Biol. 5, 76 (2017)]. Moreover, most of them have been largely conserved during metazoan evolution [L. S. Babonis, M. Q. Martindale, Philos. Trans. R. Soc. Lond. B Biol. Sci. 372, 20150477 (2017)]. How evolution has generated a vast diversity of animal morphologies with such a limited number of tools is still largely unknown. Here, we show that gain of interconnectivity between signaling pathways and the GRNs they control may have critically contributed to the origin of vertebrates. We perturbed the retinoic acid, Wnt, FGF, and Nodal signaling pathways during gastrulation in the invertebrate chordate amphioxus and zebrafish and compared the effects on gene expression and cis-regulatory elements (CREs). We found that multiple developmental genes gain response to these pathways through vertebrate-specific CREs. Moreover, in contrast to amphioxus, many of these CREs responded to multiple pathways in zebrafish, which reflects their high interconnectivity. Furthermore, we found that vertebrate-specific cell types are more enriched in highly interconnected genes than in tissues with more ancient origin. Thus, the increase of CREs in vertebrates integrating inputs from different signaling pathways probably contributed to gene expression complexity and to the formation of new cell types and morphological novelties in this lineage.This project has received funding from the European Research Council under the European Union’s Horizon 2020 research and innovation program (Grant agreement no. 740041) and the Spanish Ministerio de Economía y Competitividad (Grants BFU2016-74961-P and PID2019-103921GB-I00 to J.L.G.-S. and J.J.T.). This work was also supported by the institutional grant Unidad de Excelencia María de Maeztu (Grant MDM-2016-0687 to the Department of Gene regulation and morphogenesis of Centro Andaluz de Biología del Desarrollo). M.F. was funded by the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement [#800396]. M.S., H.E., and S.B. were supported by the CNRS, and H.E. and S.B. additionally by Agence Nationale de la Recherche (ANR) CHORELAND (Grant ANR-16-CE12-0008-01) and the Institut Universitaire de France. Y.-H.S. and J.-K.Y. are supported by intramural funds from Academia Sinica and grants from Ministry of Science and Technology, Taiwan (Grants 110-2326-B-001-006 to Y.-H.S. and 110-2621-B-001-001-MY3 to J.-K.Y.)