Implementation and validation of single-cell genomics experiments in neuroscience

Marco Colonna¹, Genevieve Konopka², Shane A Liddelow^{3

4

5

6}, Tomasz Nowakowski^{7

8

9

10}, Rajeshwar Awatramani¹¹, Helen S Bateup^{12

13

14}, Cathryn R Cadwell^{15

16

17

18}, Emre Caglayan¹⁹, Jerry L Chen^{20

21

22

23}, Jesse Gillis^{24

25}, Martin Kampmann^{26

27}, Fenna Krienen²⁸, Samuel E Marsh^{29

30

31}, Michelle Monje^{32

33}, Michael R O'Dea³⁴, Rickie Patani^{35

36}, Alex A Pollen^{37

38}, Francisco J Quintana^{39

40}, Marissa Scavuzzo^{41

42}, Matthew Schmitz³⁷, Steven A Sloan⁴³, Paul J Tesar^{41

42}, Jessica Tollkuhn²⁵, Maria Antonietta Tosches⁴⁴, Madeleine E Urbanek⁴⁵, Jonathan M Werner^{24

25}, Omer A Bayraktar⁴⁶, Ozgun Gokce^{47

48}, Naomi Habib⁴⁹

Affiliations

¹ Department of Pathology and Immunology, Washington University School of Medicine in St. Louis, St. Louis, MO, USA. mcolonna@wustl.edu.
² Department of Neuroscience, Peter O'Donnell Jr. Brain Institute, UT Southwestern Medical Center, Dallas, TX, USA. genevieve.konopka@utsouthwestern.edu.
³ Neuroscience Institute, NYU Grossman School of Medicine, New York, NY, USA. shane.liddelow@nyulangone.org.
⁴ Department of Neuroscience & Physiology, NYU Grossman School of Medicine, New York, NY, USA. shane.liddelow@nyulangone.org.
⁵ Department of Ophthalmology, NYU Grossman School of Medicine, New York, NY, USA. shane.liddelow@nyulangone.org.
⁶ Parekh Center for Interdisciplinary Neurology, NYU Grossman School of Medicine, New York, NY, USA. shane.liddelow@nyulangone.org.
⁷ Department of Neurological Surgery, University of California, San Francisco, San Francisco, CA, USA. tomasz.nowakowski@ucsf.edu.
⁸ Eli and Edythe Broad Center for Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, CA, USA. tomasz.nowakowski@ucsf.edu.
⁹ Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, USA. tomasz.nowakowski@ucsf.edu.
¹⁰ Department of Anatomy, University of California, San Francisco, San Francisco, CA, USA. tomasz.nowakowski@ucsf.edu.
¹¹ Department of Microbiology and Immunology, Northwestern University, Chicago, IL, USA.
¹² Department of Molecular and Cellular Biology, University of California, Berkeley, Berkeley, CA, USA.
¹³ Department of Neuroscience, University of California, Berkeley, Berkeley, CA, USA.
¹⁴ Chan Zuckerberg Biohub, San Francisco, CA, USA.
¹⁵ Department of Neurological Surgery, University of California, San Francisco, San Francisco, CA, USA.
¹⁶ Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, USA.
¹⁷ Department of Pathology, University of California, San Francisco, San Francisco, CA, USA.
¹⁸ Kavli Institute for Fundamental Neuroscience, University of California, San Francisco, San Francisco, CA, USA.
¹⁹ Department of Neuroscience, Peter O'Donnell Jr. Brain Institute, UT Southwestern Medical Center, Dallas, TX, USA.
²⁰ Department of Biomedical Engineering, Boston University, Boston, MA, USA.
²¹ Center for Neurophotonics, Boston University, Boston, MA, USA.
²² Department of Biology, Boston University, Boston, MA, USA.
²³ Center for Systems Neuroscience, Boston University, Boston, MA, USA.
²⁴ Department of Physiology and Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, Ontario, Canada.
²⁵ Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, USA.
²⁶ Institute for Neurodegenerative Diseases, University of California, San Francisco, San Francisco, CA, USA.
²⁷ Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA, USA.
²⁸ Princeton Neuroscience Institute, Princeton University, Princeton, NJ, USA.
²⁹ F.M. Kirby Neurobiology Center, Boston Children's Hospital, Boston, MA, USA.
³⁰ Harvard Medical School, Boston, MA, USA.
³¹ Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA, USA.
³² Department of Neurology and Neurological Sciences, Stanford University, Stanford, CA, USA.
³³ Howard Hughes Medical Institute, Stanford University, Stanford, CA, USA.
³⁴ Neuroscience Institute, NYU Grossman School of Medicine, New York, NY, USA.
³⁵ Department of Neuromuscular Disease, UCL Queen Square Institute of Neurology, London, UK.
³⁶ The Francis Crick Institute, Human Stem Cells and Neurodegeneration Laboratory, London, UK.
³⁷ Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, CA, USA.
³⁸ Department of Neurology, University of California, San Francisco, San Francisco, CA, USA.
³⁹ Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.
⁴⁰ Broad Institute of MIT and Harvard, Cambridge, MA, USA.
⁴¹ Department of Genetics and Genome Sciences, Case Western Reserve University School of Medicine, Cleveland, Ohio, OH, USA.
⁴² Institute for Glial Sciences, Case Western Reserve University School of Medicine, Cleveland, OH, USA.
⁴³ Department of Human Genetics, Emory University School of Medicine, Atlanta, GA, USA.
⁴⁴ Department of Biological Sciences, Columbia University, New York, NY, USA.
⁴⁵ Biomedical Sciences Graduate Program, University of California, San Francisco, San Francisco, CA, USA.
⁴⁶ Wellcome Sanger Institute, Cambridge, UK. ob5@sanger.ac.uk.
⁴⁷ Department of Old Age Psychiatry and Cognitive Disorders, University Hospital Bonn, Bonn, Germany. ozgun.goekce@ukbonn.de.
⁴⁸ German Center for Neurodegenerative Diseases (DZNE), Bonn, Germany. ozgun.goekce@ukbonn.de.
⁴⁹ Edmond & Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel. naomi.habib@mail.huji.ac.il.

PMID: 39627589
DOI: 10.1038/s41593-024-01814-0

Review

Implementation and validation of single-cell genomics experiments in neuroscience

Marco Colonna et al. Nat Neurosci. 2024 Dec.

. 2024 Dec;27(12):2310-2325.

doi: 10.1038/s41593-024-01814-0. Epub 2024 Dec 3.

Authors

Affiliations

¹ Department of Pathology and Immunology, Washington University School of Medicine in St. Louis, St. Louis, MO, USA. mcolonna@wustl.edu.
² Department of Neuroscience, Peter O'Donnell Jr. Brain Institute, UT Southwestern Medical Center, Dallas, TX, USA. genevieve.konopka@utsouthwestern.edu.
³ Neuroscience Institute, NYU Grossman School of Medicine, New York, NY, USA. shane.liddelow@nyulangone.org.
⁴ Department of Neuroscience & Physiology, NYU Grossman School of Medicine, New York, NY, USA. shane.liddelow@nyulangone.org.
⁵ Department of Ophthalmology, NYU Grossman School of Medicine, New York, NY, USA. shane.liddelow@nyulangone.org.
⁶ Parekh Center for Interdisciplinary Neurology, NYU Grossman School of Medicine, New York, NY, USA. shane.liddelow@nyulangone.org.
⁷ Department of Neurological Surgery, University of California, San Francisco, San Francisco, CA, USA. tomasz.nowakowski@ucsf.edu.
⁸ Eli and Edythe Broad Center for Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, CA, USA. tomasz.nowakowski@ucsf.edu.
⁹ Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, USA. tomasz.nowakowski@ucsf.edu.
¹⁰ Department of Anatomy, University of California, San Francisco, San Francisco, CA, USA. tomasz.nowakowski@ucsf.edu.
¹¹ Department of Microbiology and Immunology, Northwestern University, Chicago, IL, USA.
¹² Department of Molecular and Cellular Biology, University of California, Berkeley, Berkeley, CA, USA.
¹³ Department of Neuroscience, University of California, Berkeley, Berkeley, CA, USA.
¹⁴ Chan Zuckerberg Biohub, San Francisco, CA, USA.
¹⁵ Department of Neurological Surgery, University of California, San Francisco, San Francisco, CA, USA.
¹⁶ Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, USA.
¹⁷ Department of Pathology, University of California, San Francisco, San Francisco, CA, USA.
¹⁸ Kavli Institute for Fundamental Neuroscience, University of California, San Francisco, San Francisco, CA, USA.
¹⁹ Department of Neuroscience, Peter O'Donnell Jr. Brain Institute, UT Southwestern Medical Center, Dallas, TX, USA.
²⁰ Department of Biomedical Engineering, Boston University, Boston, MA, USA.
²¹ Center for Neurophotonics, Boston University, Boston, MA, USA.
²² Department of Biology, Boston University, Boston, MA, USA.
²³ Center for Systems Neuroscience, Boston University, Boston, MA, USA.
²⁴ Department of Physiology and Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, Ontario, Canada.
²⁵ Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, USA.
²⁶ Institute for Neurodegenerative Diseases, University of California, San Francisco, San Francisco, CA, USA.
²⁷ Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA, USA.
²⁸ Princeton Neuroscience Institute, Princeton University, Princeton, NJ, USA.
²⁹ F.M. Kirby Neurobiology Center, Boston Children's Hospital, Boston, MA, USA.
³⁰ Harvard Medical School, Boston, MA, USA.
³¹ Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA, USA.
³² Department of Neurology and Neurological Sciences, Stanford University, Stanford, CA, USA.
³³ Howard Hughes Medical Institute, Stanford University, Stanford, CA, USA.
³⁴ Neuroscience Institute, NYU Grossman School of Medicine, New York, NY, USA.
³⁵ Department of Neuromuscular Disease, UCL Queen Square Institute of Neurology, London, UK.
³⁶ The Francis Crick Institute, Human Stem Cells and Neurodegeneration Laboratory, London, UK.
³⁷ Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, CA, USA.
³⁸ Department of Neurology, University of California, San Francisco, San Francisco, CA, USA.
³⁹ Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.
⁴⁰ Broad Institute of MIT and Harvard, Cambridge, MA, USA.
⁴¹ Department of Genetics and Genome Sciences, Case Western Reserve University School of Medicine, Cleveland, Ohio, OH, USA.
⁴² Institute for Glial Sciences, Case Western Reserve University School of Medicine, Cleveland, OH, USA.
⁴³ Department of Human Genetics, Emory University School of Medicine, Atlanta, GA, USA.
⁴⁴ Department of Biological Sciences, Columbia University, New York, NY, USA.
⁴⁵ Biomedical Sciences Graduate Program, University of California, San Francisco, San Francisco, CA, USA.
⁴⁶ Wellcome Sanger Institute, Cambridge, UK. ob5@sanger.ac.uk.
⁴⁷ Department of Old Age Psychiatry and Cognitive Disorders, University Hospital Bonn, Bonn, Germany. ozgun.goekce@ukbonn.de.
⁴⁸ German Center for Neurodegenerative Diseases (DZNE), Bonn, Germany. ozgun.goekce@ukbonn.de.
⁴⁹ Edmond & Lily Safra Center for Brain Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel. naomi.habib@mail.huji.ac.il.

PMID: 39627589
DOI: 10.1038/s41593-024-01814-0

Abstract

Single-cell or single-nucleus transcriptomics is a powerful tool for identifying cell types and cell states. However, hypotheses derived from these assays, including gene expression information, require validation, and their functional relevance needs to be established. The choice of validation depends on numerous factors. Here, we present types of orthogonal and functional validation experiment to strengthen preliminary findings obtained using single-cell and single-nucleus transcriptomics as well as the challenges and limitations of these approaches.

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Conflict of interest statement

Competing interests: M.C. is a member of the Scientific Advisory Board of Vigil, NGMBio, Cartesian and Halyardtx. M.C. receives research support from Ono Pharmaceutical, is a consultant for CST and has patents pending on LILRB4 and TREM2. S.A.L. maintains a financial interest in AstronauTx and Synapticure and is on the Scientific Advisory Board of the Global BioAccess Fund. S.A.L. is an inventor on US Patents WO2018081250A1 and WO2022187517A1. M.K. is a co-scientific founder of Montara Therapeutics and serves on the Scientific Advisory Boards of Engine Biosciences, Casma Therapeutics, Cajal Neuroscience, Alector and Montara Therapeutics, and is an advisor to Modulo Bio and Recursion Therapeutics. M.K. is an inventor on US Patent 11,254,933 related to CRISPRi and CRISPRa screening, and on a US Patent Application on in vivo screening methods. All other authors declare no competing interests.

References

1. Adameyko, I. et al. Applying single-cell/nucleus genomics to studies of cellular heterogeneity and cell fate transitions in the nervous system. Nat. Neurosci. https://doi.org/10.1038/s41593-024-01827-9 (2024).
1. Bonev, B. et al. Opportunities and challenges of single-cell and spatially resolved genomics methods for neuroscience discovery. Nat. Neurosci. https://doi.org/10.1038/s41593-024-01806-0 (2024).
1. Luecken, M. D. & Theis, F. J. Current best practices in single-cell RNA-seq analysis: a tutorial. Mol. Syst. Biol. 15, e8746 (2019). - PubMed - PMC - DOI
1. Trapnell, C. Defining cell types and states with single-cell genomics. Genome Res. 25, 1491–1498 (2015). - PubMed - PMC - DOI
1. Kiselev, V. Y. et al. SC3: consensus clustering of single-cell RNA-seq data. Nat. Methods 14, 483–486 (2017). - PubMed - PMC - DOI

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Implementation and validation of single-cell genomics experiments in neuroscience

Affiliations

Implementation and validation of single-cell genomics experiments in neuroscience

Authors

Affiliations

Abstract

Conflict of interest statement

References

Publication types

MeSH terms

Grants and funding

LinkOut - more resources

Full Text Sources