CDK8 and CDK19 regulate intestinal differentiation and homeostasis via the chromatin remodeling complex SWI/SNF
Marius V. Dannappel, … , Thomas G. Boyer, Ron Firestein
Marius V. Dannappel, … , Thomas G. Boyer, Ron Firestein
Published August 25, 2022
Citation Information: J Clin Invest. 2022;132(20):e158593. https://doi.org/10.1172/JCI158593.
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Research Article Gastroenterology Genetics

CDK8 and CDK19 regulate intestinal differentiation and homeostasis via the chromatin remodeling complex SWI/SNF

Abstract

Initiation and maintenance of transcriptional states are critical for controlling normal tissue homeostasis and differentiation. The cyclin dependent kinases CDK8 and CDK19 (Mediator kinases) are regulatory components of Mediator, a highly conserved complex that orchestrates enhancer-mediated transcriptional output. While Mediator kinases have been implicated in the transcription of genes necessary for development and growth, its function in mammals has not been well defined. Using genetically defined models and pharmacological inhibitors, we showed that CDK8 and CDK19 function in a redundant manner to regulate intestinal lineage specification in humans and mice. The Mediator kinase module bound and phosphorylated key components of the chromatin remodeling complex switch/sucrose non-fermentable (SWI/SNF) in intestinal epithelial cells. Concomitantly, SWI/SNF and MED12-Mediator colocalized at distinct lineage-specifying enhancers in a CDK8/19–dependent manner. Thus, these studies reveal a transcriptional mechanism of intestinal cell specification, coordinated by the interaction between the chromatin remodeling complex SWI/SNF and Mediator kinase.

Authors

Marius V. Dannappel, Danxi Zhu, Xin Sun, Hui Kheng Chua, Marle Poppelaars, Monica Suehiro, Subash Khadka, Terry C.C. Lim Kam Sian, Dhanya Sooraj, Melissa Loi, Hugh Gao, Daniel Croagh, Roger J. Daly, Pouya Faridi, Thomas G. Boyer, Ron Firestein

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Figure 2

CDK8/19 kinase activity is necessary for intestinal epithelial cell intrinsic growth and differentiation.

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CDK8/19 kinase activity is necessary for intestinal epithelial cell intr...
(A) Schematic representation of the genetic small intestinal organoid model. (B) qRT-PCR analysis of Cdk8 and Cdk19 gene expression from small intestinal organoids with the indicated genotypes 1 week after tamoxifen treatment. Representative data from 2 independent experiments. Holm-Šidák multiple-comparison test. (C) Immunoblot of small intestinal organoids from VillinCreERT2, VillinCreERT2/Cdk8fl/fl, and VillinCreERT2/Cdk8fl/fl/Cdk19–/– mice 1 week after 4-OHT treatment for 24 hours. (D) Top: Relative organoid numbers (to P1) of lines with the indicated genotypes for 4 passages after treatment with EtOH or 4-OHT. Bottom: Representative images of VillinCreERT2, CDK8iIEC-KO, Cdk8fl/fl/Cdk19–/–, and CDK8iIEC-KO/Cdk19–/– organoids at the end of passage 2 after 4-OHT treatment. Representative data from 3 independent experiments. Two-way ANOVA test (Šidák’s multiple-comparison test). Scale bars: 500 μm. (E) Relative numbers of EtOH- and 4-OHT–treated VillinCreERT2/Cdk8fl/fl/Cdk19D173A/– organoids for 4 passages. Representative data from 3 independent experiments. Unpaired 2-tailed t test. (F) Representative images of VillinCreERT2/Cdk8fl/fl/Cdk19D173A/– organoids at the end of passage 2 after treatment with EtOH or 4-OHT. Scale bars: 200 μm. (G) VillinCreERT2/Cdk8fl/fl/Cdk19D173A/– organoid viability at passage 2 after EtOH and 4-OHT treatment measured with alamarBlue HS (Thermo Fisher Scientific). Representative of 3 independent experiments. Unpaired 2-tailed t test. (H) Relative organoid numbers of intestinal organoids (C57BL/6 WT) treated with the indicated concentrations of Cp32 for 4 passages. Representative data from 3 independent experiments. Ordinary 1-way ANOVA. (I) Representative images of C57BL/6 small intestinal organoids treated with DMSO or 1 μM Cp32 at the end of passage 2. Scale bars: 500 μm. (J) Bar graph shows human small intestinal organoid viability (alamarBlue HS) 96 hours after transduction with shRNA targeting CCNC or nontargeting control. Unpaired 2-tailed t test. Data represent mean ± SEM. **P ≤ 0.01, ***P ≤ 0.005, ****P ≤ 0.001.