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Principal Investigator: Matthias Stadtfeld
Organization: WEILL MEDICAL COLL OF CORNELL UNIV
Fiscal Year: 2024
Award: $169,500
Funding agency: National Institute of Neurological Disorders and Stroke
Project Summary
Mutations affecting the activity or expression of cell type-specific DNA-binding transcription factors (TFs) are a
common cause for a range of human diseases and congenital disorders. Despite the established relevance of
TF function for human health, the inaccessibility of the developing embryo and the complexity of tissue patterning
in vivo has resulted in significant gaps in our basic molecular understanding on how mutations in TFs result in
organ-specific pathologies. An example for this is haploinsufficiency of OTX2 – an essential homeobox TF
expressed in pluripotent tissues of the early embryo and throughout nervous system development – which results
in several rare, poorly understood congenital disorders that affect neuronal, sensory, and endocrine components
of the central and peripheral nervous system (NS). An assessment of clinical case reports and the published
literature on OTX2 function in animal models let us to hypothesize that NS-specific pathologies downstream of
OTX2 haploinsufficiency are caused by dysregulation of gene loci controlled by neurodevelopmental cis-
regulatory elements and chromatin-associated protein complexes that are uniquely sensitive to OTX2 dosage.
Our long-term goal is to understand how molecular and cellular pathologies caused by OTX2 deficiency in the
nervous system can be effectively reversed. The objective of this proposal is to implement a tractable
experimental platform that combines custom modifications of the endogenous OTX2 locus with the directed
differentiation of human pluripotent stem cells (hPSCs) to identify gene regulatory mechanisms employed by this
TF during NS development. To allow discerning gene regulatory elements such as distal enhancers and
associated gene loci controlled by OTX2 during human NS development we have engineered human pluripotent
stem cells to harbor a multipurpose degron allele for rapid depletion and efficient immunoprecipitation of
endogenous OTX2, which we will leverage to identify OTX2-dependent enhancers during early stages of ex vivo
human NS specification (Aim 1). In parallel, we will engineer a second multipurpose allele that will allow
identification of OTX2 interaction partners by proximity ligation and utilize it to discern OTX2 containing gene
regulatory protein complexes active in hPSCs and derivative NS (Aim 2). Together, our proposed experiments
will provide a workable molecular framework for OTX2-dependent gene regulatory networks in the developing
human nervous system, thereby helping to close important gaps in our understanding of OTX2 function in
physiological and pathological conditions.
Terms: <Acute><Address><Affect><Alleles><Allelomorphs><Animal Model><Animal Models and Related Studies><Antigenic Determinants><Basal Transcription Factor><Basal transcription factor genes><Binding Determinants><Bio-Informatics><Bioinformatics><Biological><Body System><Body Tissues><CNS Nervous System><Case Study><Central Nervous System><Cephalic><ChIP assay><Chromatin><Chromatin Remodeling Complex><Chromatin Remodeling Factor><Chromosomal, Gene, or Protein Abnormality><Clinical><Closure by Ligation><Complex><Congenital Disorders><Cranial><Custom><Cytogenetic or Molecular Genetic Abnormality><DNA Binding><DNA Binding Interaction><DNA bound><Development><Disease><Disorder><Distal><Ectoderm><Embryo><Embryonic><Endocrine><Engineering><Enhancer Elements><Enhancers><Epitopes><Expression Signature><Future><Gene Expression><Gene Expression Monitoring><Gene Expression Pattern Analysis><Gene Expression Profile><Gene Expression Profiling><Gene Transcription><General Transcription Factor Gene><General Transcription Factors><Genes><Genetic Abnormality><Genetic Alteration><Genetic Change><Genetic Engineering><Genetic Engineering Biotechnology><Genetic Engineering Molecular Biology><Genetic Enhancer Element><Genetic Transcription><Genetic defect><Glia><Glial Cells><Goals><Health><Homeo Boxes><Homeobox><Human><Human Development><Human Engineering><Immune Precipitation><Immunoprecipitation><Influenza HA><Influenza Hemagglutinin><KI mice><KO mice><Knock-in Mouse><Knock-out Mice><Knockout Mice><Kolliker's reticulum><Label><Ligation><Literature><Microphthalmos><Modeling><Modern Man><Modification><Molecular><Molecular Abnormality><Mutation><Nature><Nerve Cells><Nerve Unit><Nervous System><Nervous System Diseases><Nervous System Disorder><Neural Cell><Neuraxis><Neurocyte><Neuroglia><Neuroglial Cells><Neurologic Body System><Neurologic Disorders><Neurologic Organ System><Neurological Disorders><Neurons><Non-neuronal cell><Nonneuronal cell><Null Mouse><Organ><Organ System><Orphan Disease><Pathologic><Pathology><Patients><Pattern><Peripheral Nervous System><Phenotype><Physiologic><Physiological><Pituitary Hormones><Property><Proteins><Proteomics><Publishing><RANK protein><RNA Expression><Rare Diseases><Rare Disorder><Recombinant DNA Technology><Regulatory Element><Regulatory Protein><Resolution><Retinal Dystrophy><Role><Sensory><Series><Specific qualifier value><Specified><TNFRSF11A gene product><Technology><Therapeutic><Therapeutic Intervention><Tissues><Transcript Expression Analyses><Transcript Expression Analysis><Transcription><Transcription Factor Proto-Oncogene><Transcription factor genes><Transgenic Organisms><Visualization><Work><analyze gene expression><biologic><candidate identification><case report><cell type><cellular pathology><chromatin immunoprecipitation><chromatin modifier><customs><design><designing><developmental><directed differentiation><dosage><enhancer sequence><epigenome profiling><epigenomic profiling><experiment><experimental research><experimental study><experiments><flu HA><flu hemagglutinin><gene expression analysis><gene expression assay><gene expression pattern><gene expression signature><gene locus><gene regulatory network><genetic enhancer sequence><genetic locus><genetic regulatory protein><genetically engineered><genome mutation><genomic location><genomic locus><hESC><hormone deficiency><human ES cell><human ESC><human disease><human embryonic stem cell><human pluripotent stem cell><in vivo><influenza viral HA><influenza viral hemagglutinin><influenza virus HA><influenza virus hemagglutinin><insight><intervention therapy><knockin mice><loss of function mutation><microphthalmia><model of animal><molecular aberrations><molecular pathology><nanophthalmos><nerve cement><nervous system development><neurological disease><neuronal><novel><orphan disorder><protein complex><receptor activator of NF-kappaB><regulatory gene product><resolutions><social role><targeted drug therapy><targeted drug treatments><targeted therapeutic><targeted therapeutic agents><targeted therapy><targeted treatment><therapeutic agent development><therapeutic development><tool><transcription factor><transcriptional profile><transcriptional profiling><transcriptional signature><transgenic>