CRISPRi screens in human iPSC-derived astrocytes elucidate regulators of distinct inflammatory reactive states

Kun Leng; Indigo V L Rose; Hyosung Kim; Wenlong Xia; Wilber Romero-Fernandez; Brendan Rooney; Mark Koontz; Emmy Li; Yan Ao; Shinong Wang; Mitchell Krawczyk; Julia Tcw; Alison Goate; Ye Zhang; Erik M Ullian; Michael V Sofroniew; Stephen P J Fancy; Matthew S Schrag; Ethan S Lippmann; Martin Kampmann

doi:10.1038/s41593-022-01180-9

CRISPRi screens in human iPSC-derived astrocytes elucidate regulators of distinct inflammatory reactive states

Nat Neurosci. 2022 Nov;25(11):1528-1542. doi: 10.1038/s41593-022-01180-9. Epub 2022 Oct 27.

Authors

Kun Leng^{1

2

3}, Indigo V L Rose^{4

5}, Hyosung Kim⁶, Wenlong Xia⁷, Wilber Romero-Fernandez⁸, Brendan Rooney⁴, Mark Koontz⁹, Emmy Li^{4

10}, Yan Ao¹¹, Shinong Wang¹¹, Mitchell Krawczyk^{12

13}, Julia Tcw^{14

15

16}, Alison Goate^{15

16

17}, Ye Zhang¹³, Erik M Ullian⁹, Michael V Sofroniew¹¹, Stephen P J Fancy⁷, Matthew S Schrag^{8

18

19}, Ethan S Lippmann^{6

8

18

19

20}, Martin Kampmann^{21

22

23}

Affiliations

¹ Institute for Neurodegenerative Diseases, University of California, San Francisco, San Francisco, CA, USA. kun.leng@ucsf.edu.
² Biomedical Sciences Graduate Program, University of California, San Francisco, San Francisco, CA, USA. kun.leng@ucsf.edu.
³ Medical Scientist Training Program, University of California, San Francisco, San Francisco, CA, USA. kun.leng@ucsf.edu.
⁴ Institute for Neurodegenerative Diseases, University of California, San Francisco, San Francisco, CA, USA.
⁵ Neuroscience Graduate Program, University of California, San Francisco, San Francisco, CA, USA.
⁶ Department of Chemical and Biomolecular Engineering, Vanderbilt University, Nashville, TN, USA.
⁷ Departments of Neurology and Pediatrics, School of Medicine, University of California, San Francisco, San Francisco, CA, USA.
⁸ Department of Neurology, Vanderbilt University Medical Center, Nashville, TN, USA.
⁹ Department of Ophthalmology, School of Medicine, University of California, San Francisco, San Francisco, CA, USA.
¹⁰ Biomedical Sciences Graduate Program, University of California, San Francisco, San Francisco, CA, USA.
¹¹ Department of Neurobiology, University of California, Los Angeles, Los Angeles, CA, USA.
¹² Interdepartmental PhD Program in Neuroscience, University of California, Los Angeles, Los Angeles, CA, USA.
¹³ Department of Psychiatry and Biobehavioral Sciences, David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, CA, USA.
¹⁴ Department of Pharmacology and Experimental Therapeutics, Boston University School of Medicine, Boston, MA, USA.
¹⁵ Nash Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
¹⁶ Ronald M. Loeb Center for Alzheimer's Disease, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
¹⁷ Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
¹⁸ Vanderbilt Memory and Alzheimer's Center, Vanderbilt University Medical Center, Nashville, TN, USA.
¹⁹ Vanderbilt Brain Institute, Vanderbilt University, Nashville, TN, USA.
²⁰ Department of Biomedical Engineering, Vanderbilt University, Nashville, TN, USA.
²¹ Institute for Neurodegenerative Diseases, University of California, San Francisco, San Francisco, CA, USA. martin.kampmann@ucsf.edu.
²² Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, USA. martin.kampmann@ucsf.edu.
²³ Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA, USA. martin.kampmann@ucsf.edu.

Abstract

Astrocytes become reactive in response to insults to the central nervous system by adopting context-specific cellular signatures and outputs, but a systematic understanding of the underlying molecular mechanisms is lacking. In this study, we developed CRISPR interference screening in human induced pluripotent stem cell-derived astrocytes coupled to single-cell transcriptomics to systematically interrogate cytokine-induced inflammatory astrocyte reactivity. We found that autocrine-paracrine IL-6 and interferon signaling downstream of canonical NF-κB activation drove two distinct inflammatory reactive signatures, one promoted by STAT3 and the other inhibited by STAT3. These signatures overlapped with those observed in other experimental contexts, including mouse models, and their markers were upregulated in human brains in Alzheimer's disease and hypoxic-ischemic encephalopathy. Furthermore, we validated that markers of these signatures were regulated by STAT3 in vivo using a mouse model of neuroinflammation. These results and the platform that we established have the potential to guide the development of therapeutics to selectively modulate different aspects of inflammatory astrocyte reactivity.

Publication types

Research Support, Non-U.S. Gov't
Research Support, N.I.H., Extramural

MeSH terms

Alzheimer Disease*
Astrocytes
Cytokines
Humans
Induced Pluripotent Stem Cells*
Inflammation
Signal Transduction

Substances

Cytokines

Abstract

Publication types

MeSH terms

Substances

Grants and funding