1. Academic Validation
  2. Identification of astrocyte regulators by nucleic acid cytometry

Identification of astrocyte regulators by nucleic acid cytometry

  • Nature. 2023 Jan 4. doi: 10.1038/s41586-022-05613-0.
Iain C Clark # 1 2 3 Michael A Wheeler # 1 4 Hong-Gyun Lee 1 Zhaorong Li 1 4 Liliana M Sanmarco 1 Shravan Thaploo 1 Carolina M Polonio 1 Seung Won Shin 3 Giulia Scalisi 1 Amy R Henry 5 Joseph M Rone 1 Federico Giovannoni 1 Marc Charabati 1 Camilo Faust Akl 1 Dulce M Aleman 1 Stephanie E J Zandee 6 Alexandre Prat 6 Daniel C Douek 5 Eli A Boritz 5 Francisco J Quintana 7 8 Adam R Abate 9
Affiliations

Affiliations

  • 1 Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, 02115, USA.
  • 2 Department of Bioengineering and Therapeutic Sciences, School of Pharmacy, University of California San Francisco, San Francisco, CA, 94158, USA.
  • 3 Department of Bioengineering, College of Engineering, California Institute for Quantitative Biosciences, QB3, University of California Berkeley, Berkeley, CA, 94720, USA.
  • 4 Broad Institute of MIT and Harvard, Cambridge, MA, 02142, USA.
  • 5 Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, 20892, USA.
  • 6 Neuroimmunology Research Lab, Centre de Recherche du Centre Hospitalier de l'Université de Montréal (CRCHUM), Montreal, QC, H2X 0A9, Canada.
  • 7 Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, 02115, USA. [email protected]
  • 8 Broad Institute of MIT and Harvard, Cambridge, MA, 02142, USA. [email protected]
  • 9 Department of Bioengineering and Therapeutic Sciences, School of Pharmacy, University of California San Francisco, San Francisco, CA, 94158, USA. [email protected]
  • # Contributed equally.
Abstract

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS)1. Astrocytes are heterogeneous CNS-resident glial cells that participate in the pathogenesis of MS and its model experimental autoimmune encephalomyelitis (EAE)2,3. However, few unique surface markers are available for the isolation of astrocyte subsets, preventing their analysis and the identification of candidate therapeutic targets; these limitations are further amplified by the rarity of pathogenic astrocytes. To address these challenges, we developed FIND-seq (Focused Interrogation of cells by Nucleic acid Detection and Sequencing), a high-throughput microfluidic cytometry method that combines encapsulation of cells in droplets, PCR-based detection of target nucleic acids, and droplet sorting to enable in-depth transcriptomic analyses of cells of interest at single-cell resolution. We applied FIND-seq to study the regulation of astrocytes characterized by the splicing-driven activation of the transcription factor XBP1, which promotes disease pathology in MS and EAE4. Using FIND-seq in combination with conditional knock-out mice, in vivo CRISPR/Cas9-driven genetic perturbation studies, and bulk and single-cell RNA-seq analyses of mouse EAE and human MS samples, we identified a new role for the nuclear receptor NR3C2 and its corepressor NCOR2 in limiting XBP1-driven pathogenic astrocyte responses. In summary, FIND-seq enabled the identification of a therapeutically targetable mechanism that limits XBP1-driven pathogenic astrocyte responses. FIND-seq allows the investigation of previously inaccessible cells, including rare cell subsets defined by unique gene expression signatures or other nucleic acid markers.

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