Small-molecule screen reveals pathways that regulate C4 secretion in stem cell-derived astrocytes

Francesca Rapino; Ted Natoli; Francesco Limone; Erin O'Connor; Jack Blank; Matthew Tegtmeyer; William Chen; Erika Norabuena; Juhi Narula; Dane Hazelbaker; Gabriella Angelini; Lindy Barrett; Alison O'Neil; Ursula K Beattie; Jessica M Thanos; Heather de Rivera; Steven D Sheridan; Roy H Perlis; Steven A McCarroll; Beth Stevens; Aravind Subramanian; Ralda Nehme; Lee L Rubin

doi:10.1016/j.stemcr.2022.11.018

Small-molecule screen reveals pathways that regulate C4 secretion in stem cell-derived astrocytes

Stem Cell Reports. 2023 Jan 10;18(1):237-253. doi: 10.1016/j.stemcr.2022.11.018. Epub 2022 Dec 22.

Authors

Francesca Rapino¹, Ted Natoli², Francesco Limone³, Erin O'Connor⁴, Jack Blank⁴, Matthew Tegtmeyer⁵, William Chen⁴, Erika Norabuena⁴, Juhi Narula⁴, Dane Hazelbaker⁵, Gabriella Angelini⁵, Lindy Barrett⁵, Alison O'Neil⁴, Ursula K Beattie⁴, Jessica M Thanos⁶, Heather de Rivera⁷, Steven D Sheridan⁶, Roy H Perlis⁶, Steven A McCarroll⁷, Beth Stevens⁸, Aravind Subramanian², Ralda Nehme⁵, Lee L Rubin⁹

Affiliations

¹ Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, Cambridge, MA 02138, USA; Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA. Electronic address: francesca_rapino@harvard.edu.
² Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
³ Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, Cambridge, MA 02138, USA; Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Leiden University Medical Center, LUMC, 2333 ZA Leiden, the Netherlands.
⁴ Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, Cambridge, MA 02138, USA.
⁵ Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA; Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
⁶ Center for Quantitative Health, Center for Genomic Medicine and Department of Psychiatry, Massachusetts General Hospital, Boston, MA, USA; Department of Psychiatry, Harvard Medical School, Boston, MA, USA.
⁷ Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Department of Genetics, Harvard Medical School, Boston, MA, USA.
⁸ Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Department of Neurology, F.M. Kirby Neurobiology Center, Boston Children's Hospital, Harvard Medical School, Boston, MA, USA.
⁹ Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, Cambridge, MA 02138, USA; Stanley Center for Psychiatric Research, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA. Electronic address: lee_rubin@harvard.edu.

Abstract

In the brain, the complement system plays a crucial role in the immune response and in synaptic elimination during normal development and disease. Here, we sought to identify pathways that modulate the production of complement component 4 (C4), recently associated with an increased risk of schizophrenia. To design a disease-relevant assay, we first developed a rapid and robust 3D protocol capable of producing large numbers of astrocytes from pluripotent cells. Transcriptional profiling of these astrocytes confirmed the homogeneity of this population of dorsal fetal-like astrocytes. Using a novel ELISA-based small-molecule screen, we identified epigenetic regulators, as well as inhibitors of intracellular signaling pathways, able to modulate C4 secretion from astrocytes. We then built a connectivity map to predict and validate additional key regulatory pathways, including one involving c-Jun-kinase. This work provides a foundation for developing therapies for CNS diseases involving the complement cascade.

Keywords: C4; CMap; ELISA-based screen; ESCs; L1000; astrocytes; complement component C4; complement system; connectivity map; disease modeling; human embryonic stem cells; human induced pluripotent stem cells; iPSCs; screen; stem cell-derived astrocytes.

Small-molecule screen reveals pathways that regulate C4 secretion in stem cell-derived astrocytes

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