Joel Glover group

Background
The use of stem cells to produce neurons for treating neurological diseases depends on a deep understanding of how neurons differentiate during embryonic and fetal life. Research in our laboratory is focused on two main topics: 1) the regional patterning and differentiation of neurons and the development of functional connections between them that give rise to defined sensorimotor circuits, and 2) neuronal differentiation of human adult somatic stem cells and of human pluripotent stem cells, as a means of generating the neurons that make up these circuits.  The research group consists of 15 scientists, postdocs, PhD students and technicians. Our technical expertise lies in neuronal tract tracing, the characterization of neuronal differentiation at the molecular (mRNA and protein) and structural level, the use of electrophysiological and optical recording techniques to assess functional synaptic connectivity, and more recently in vivo MRI-based tracking of human stem cells in animal models.

 

Research goals
Our long term goal is to understand how specific sensorimotor circuits in the brain stem and spinal cord develop and how stem cells can be used both in potential replacement therapies for such conditions as spinal cord injury and as tools to better understand the molecular and cellular mechanisms underlying neuronal patterning, differentiation and synaptic coupling. We are focusing in particular on the neuronal circuits that mediate the descending control of spinal motor programs and the ascending control of eye movements, including vestibulospinal, reticulospinal and vestibulo-ocular projections.

  

Projects

  • Development of mammalian and avian reticulospinal, vestibulospinal and vestibulo-ocular projections, including molecular characterization of projection neuron subpopulations and optical and electrophysiological recording of synaptic activation patterns.
  • Anteroposterior and dorsoventral patterning of reticular and vestibular projection neurons, through fate-mapping and correlation with AP and DV patterning gene expression.
  • Molecular, anatomical and physiological development of extraocular motoneurons.
    Characterization of the molecular mechanisms underlying regulative regeneration in the embryonic spinal cord, wherein neural tissue spontaneously repairs itself through controlled proliferation and differentiation.
  • Generation of neurons from human embryonic and adult somatic stem cells as a platform for developing regenerative treatments for spinal cord injury.

 

  Achievements

  • First demonstration of the requirement of serotonin for animal locomotion (Glover and Kramer, Science, 1984).
  • Invented the technique of retrograde labeling using conjugated dextrans, now used world-wide (Glover et al, J Neurosci Methods, 1986). 
  • First demonstration of clonal relationships giving rise to neuron columns in a laminar brain structure, and the progression from symmetric to asymmetric divisions in clones of neural stem cells (Gray et al, PNAS, 1988).
  • First description of the development of an identified spinal interneuron in an amniote embryo (Eide and Glover, J Neurosci, 1996).
  • Demonstration of a retinoid-dependent segmental patterning of sympathetic preganglionic neurons (Forehand et al., PNAS, 1998).
  • Segmental analyses of reticulospinal and vestibular projection neurons in avians and mammals (Glover and Petursdottir, J Neurobiol, 1991; Diaz et al, Dev Biol 1998; Auclair et al, J Comp Neurol, 1999; Pasqualetti et al, J Neurosci, 2007).
  • First demonstration of an in vivo differentiation of adult human non-neural somatic stem cells to neurons (Sigurjonsson et al, PNAS, 2005).
  • First characterization using dynamic imaging methods of the emergence of an entire polysynaptic reflex pathway in a vertebrate (Glover et al, submitted).

 

International collaborations

Filippo Rijli, Friedrich Miescher Institute, Switzerland
Sascha du Lac, Salk Institute, USA
Carmina Díaz, Universidad de Albacete, Spain
Susan Dymecki, Harvard University, USA
Jeff Lichtman, Harvard University, USA
Estelle Gauda, Johns Hopkins University, USA
Bernd Fritzsch, University of Iowa, USA
Katsushige Sato, Komazawa Women's University, Japan
Yoko Momose-Sato, Kanto Gakuin University College, Japan

 

 

Selected publications

Szokol K, Glover JC, Perreault MC (2011) Organization of functional synaptic connections between medullary reticulospinal neurons and lumbar descending commissural interneurons in the neonatal mouse. J Neurosci 31:4731-4742.

Boulland JL, Halasi G, Kasumacic N, Glover JC (2010) Xenotransplantation of human stem cells into the chicken embryo. J Vis Exp, Jul 11;(41). pii: 2071. doi: 10.3791/2071.

Kasumacic N, Glover JC, Perreault MC (2010) Segmental patterns of vestibular-mediated synaptic inputs to axial and limb motoneurons in the neonatal mouse assessed by optical recording. J Physiol 588:4905-4925. 

Szokol K, Glover JC, Perreault MC (2008) Differential origin of reticulospinal drive to motoneurons innervating trunk and hindlimb muscles in the mouse revealed by optical recording. J Physiol 586:5259-5276. 

Pasqualetti M, Díaz C, Renaud JS, Rijli F and Glover JC (2007) Fate-mapping the mammalian hindbrain: Segmental origins of vestibular projection neurons assessed using rhombomere-specific Hoxa2 enhancer elements in the mouse embryo. J Neurosci 27:9670-9681.

Sigurjonsson OE, Perreault MC, Egeland T, Glover JC (2005) Adult human hematopoietic stem cells produce neurons efficiently in the regenerating chicken embryo spinal cord. PNAS 102:5227-5232. 

Glover JC (2000) The development of specific connectivity between premotor neurons and motoneurons in the brain stem and spinal cord. Physiological Reviews 80:615-647. 

Auclair F, Marchand R, Glover JC (1999) Regional patterning of reticulospinal and vestibulospinal neurons in the hindbrain of rat and mouse embryos. J Comp Neurol 411: 288-300. 

Forehand CF, Ezerman EB, Goldblatt J, Glover JC (1998) The segment-specific pattern of sympathetic preganglionic projections is altered by retinoids. PNAS 95:10878-10883. 

Eide AL, Glover JC (1996) The development of an identified spinal commissural interneuron population in an amniote: the neurons of the avian Hofmann nuclei. J Neurosci 16:5749-5761. 

Glover JC, Petursdottir G (1991) Regional specificity of developing reticulospinal, vestibulospinal, and vestibulo-ocular projections in the chicken embryo. J Neurobiol 22:353-376. 

Gray GE, Glover JC, Majors J, Sanes JR (1988) Radial arrangement of clonally related cells in the chicken optic tectum: Lineage analysis with a recombinant retrovirus. PNAS 85:7356-7360. 

Glover JC, Petursdottir G, Jansen JKS (1986) Fluorescent dextran-amines used as axonal tracers in the nervous system of the chicken embryo. J Neurosci Methods 18: 243-254.

 

Philippe Collas group

 

Chromatin and nuclear architecture in stem cells

The CollasLab investigates principles of 3-dimensional genome architecture which pattern lineage-specific stem cell differentiation in health and disease contexts.
See also www.collaslab.org.

 

Research goal

The 3D layout of chromatin plays important roles in the establishment of gene expression programs governing cell fate decisions.We are addressing three main questions:

  • How is 3D genome conformation regulated during fat stem cell differentiation?
  • How do laminopathy-causing lamin mutations affect chromatin conformation?
  • How do histone H3 variants contribute to chromatin homeostasis in normal and cancer cells?

Our work combines cell biological, genomics and genome structure modeling approaches using patient material and engineered stem cells.

 

The lab’s research history in brief

  • disassembly and reformation of the nuclear envelope (Steen 2000 J Cell Biol; Steen 2001 J Cell Biol; Martins 2003 J Cell Biol)
  • cell and nuclear reprogramming (Håkelien 2002 Nature Biotech; Taranger 2005 Mol Biol Cell; Freberg 2007 Mol Biol Cell)
  • chromatin immunoprecipitation (ChIP) assays for small cell numbers (Dahl 2008 Nature Protoc; 2009 Genome Biol)
  • epigenetic patterning of developmental gene expression (Lindeman 2011 Dev Cell; Andersen 2012 Genome Biol) and of adipose stem cell differentiation (Boquest 2007 Stem Cells; Sørensen 2010 Mol Biol Cell; Shah 2014 BMC Genomics; Rønningen 2015 BBRC)
  • nuclear lamin - chromatin interactions during adipogenic differentiation (Lund 2013 Genome Res; Lund 2014 Nucl Acids Res; Oldenburg 2014 Hum Mol Genet; Rønningen 2015 Genome Res)
  • deposition of histone H3 variants into chromatin (Delbarre 2010 Mol Biol Cell; Delbarre 2013 Genome Res; Ivanauskiene 2014 Genome Res; Delbarre 2017 Genome Res)
  • 3D genome modeling (Paulsen 2015 PloS Comput Biol; Sekelja 2016 Genome Biol; Paulsen 2017 Genome Biol)

 

Projects:

 

3D organization of the stem cell genome

3D genome organization of the genome influences cell- and time-specific blueprints of gene expression. Some aspects of 3D genome conformation vary between cell types, suggesting developmental regulation. 3D genome conformation entails interactions between chromosomes, forming interactions hubs called topologically-associating domains (TADs). At the nuclear periphery, chromosomes interact with the nuclear lamina through lamin-associated domains (LADs). These interactions are dynamic during differentiation. We are investigating links between cellular metabolism and changes in spatial genome conformation during differentiation of adipose stem cells in normal and pathological conditions.

Ongoing research:

  • Computational methods for 3D and 4D modeling of genome architecture
  • Functional relationships between 3D chromatin folding, nuclear envelope-chromatin interactions, epigenetic states and lineage-specific differentiation capacity

 

Nuclear lamina, genome organization & adipose stem cell function

The nuclear envelope regulates gene expression by interacting with chromatin. It consists of a double nuclear membrane, nuclear pores and the nuclear lamina, a meshwork of A-type lamins (LMNA, LMNC) and B-type lamins (LMNB1, LMNB2). Mutations in LMNA cause laminopathies, which include progeria, muscle dystrophies and partial lipodystrophies. Familial partial lipodystrophy of Dunnigan type (FPLD2) mainly affects adipose tissue and leads to severe metabolic disorders. We use patient cells, patient-derived iPS cells and engineered adipose stem cells to address the impact of lipodystrophic LMNA mutations on differentiation of adipose stem cells (ASCs), formation and composition of LADs and 3D genome conformation.

Ongoing research:

  • Identification of determinants of nuclear envelope-chromatin interactions during lineage-specific differentiation and in laminopathy contexts
  • Bioinformatics development

 

Histone variants and chromatin homeostasis

A class of pediatric gliomas (diffuse intrinsic pontine gliomas, DIPGs), is driven by mutations in the H3F3A gene encoding histone variant H3.3. The most prominent DIPG driver H3.3 mutation is H3.3K27M, which results in global reduction of H3K27me3. Other H3.3 mutations include H3.3G34R, which affects other nearby H3 modifications.

Ongoing research:

  • Mechanisms of deposition of histone H3 variants into chromatin
  • Role of H3.3 on  chromatin homeostasis
  • Impact of H3.3 mutations on chromatin and nuclear architecture in glioblastomas

 

Key collaborations

  • Louis Casteilla, StromaLab, University of Toulouse, France (adipose tissue metabolism)
  • Jacques Grill, Institut Gustave Roussy, Villejuif, France (H3.3 mutations and pediatric glioblastoma)
  • Anna Kostareva, Almazov Research Center, St. Petersburg, Russia (mesenchymal stem cells)
  • Stefan Pfister, DKFZ, Heidelberg, Germany (pediatric glioblastoma)
  • Corinne Vigouroux, Hôpital Saint Antoine, INSERM, Paris, France (lipodystrophies)
  • Lee Wong, Monash University, Clayton, Australia (heterochromatin)
  • David Tremethick, John Curtis School of Medical Research, Australian National University, Canberra, Australia (genome conformation)

  

Some recent publications

Delbarre, E, Ivanauskiene K, Spirkoski J, Shah A, Vekterud K, Moskaug JØ, Bøe SO, Wong L, Küntziger T, Collas P. 2017. PML protein organizes heterochromatin domains where it regulates histone H3.3 deposition by ATRX/DAXX. Genome Res. In press.

Paulsen J, Sekelja M, Oldenburg AR, Barateau A, Briand N, Delbarre E, Shah, A, Sørensen AL, Vigouroux C, Buendia B, Collas P. 2017. Chrom3D: three-dimensional genome modeling from Hi-C and nuclear lamin-genome contacts. Genome Biol. 18, 21-29.

Sekelja M., Paulsen J., Collas P. 2016. 4D nucleomes in single cells: what can computational modeling reveal about spatial chromatin conformation? Genome Biol 17, 54

Rønningen T., Shah A, Oldenburg AR, Vekterud K, Delbarre E, Moskaug JØ, Collas P. 2015. Prepatterning of differentiation-driven nuclear lamin A/C-associated chromatin domains by GlcNAcylated histone H2B. Genome Res 25, 1825-1835.

Ivanauskiené K., Delbarre E., McGhie J.D., Küntziger T., Wong L.H., Collas P. 2014. The PML-associated protein DEK regulates the balance of H3.3 loading on chromatin and is important for for telomere integrity. Genome Res. 24, 1584-1594

 

 

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