citations

IRF2 is a master regulator of human keratinocyte stem cell fate

Authors:

Mercado N, Schutzius G, Kolter C, Estoppey D, Bergling S, Roma G, Gubser Keller C, Nigsch F, Salathe A, Terranova R, Reece-Hoyes J, Alford J, Russ C, Knehr J, Hoepfner D, Aebi A, Ruffner H, Beck TC, Jagannathan S, Olson CM, Sheppard HE, Elsarrag SZ, Bouwmeester T, Frederiksen M, Lohmann F, Lin CY, Kirkland S.

In:

Source: Nat Commun.
Publication Date: (2019)
Issue: 10: 4676

Research Area:

Stem Cells

Experiment

Human dermo-epidermal 3D models.

Human primary dermal fibroblasts (CELLnTEC, Bern, Switzerland, Lot#ES1303222) were expanded in DMEM/F-12 with GlutaMAX™ supplemented with 10% fetal calf serum

(ThermoFisher Scientific, Zug, Switzerland) up to passage 6 before used in the dermo-epidermal model.

The dermal part was prepared with the RAFT kit (LONZA, Visp, Switzerland, cat#016-0R94) according to the manufacturer’s instructions. The cell-collagen

solution with a final cell density of 4 × 104 cells per well was cast into a HTS Transwell®-24-well Permeable Support with 0.4 µm Pore Polyester Membrane and

6.5mm insert diameter (Corning, cat#3378). RAFT absorbers (LONZA, cat#016- 1R33) were placed on top for 15 min to condense the cell–collagen mix. The cells

were left to mature for 1 week at 37 °C, 5% CO2 and medium (DMEM/F-12 with GlutaMAX™ supplemented with 5% fetal calf serum and freshly supplemented with

284 µM L-ascorbic acid (Sigma-Aldrich) was changed every 2–3 days.

Abstract

Resident adult epithelial stem cells maintain tissue homeostasis by balancing self-renewal and differentiation. The stem cell potential of human epidermal keratinocytes is retained in vitro but lost over time suggesting extrinsic and intrinsic regulation. Transcription factor-controlled regulatory circuitries govern cell identity, are sufficient to induce pluripotency and transdifferentiate cells. We investigate whether transcriptional circuitry also governs phenotypic changes within a given cell type by comparing human primary keratinocytes with intrinsically high versus low stem cell potential. Using integrated chromatin and transcriptional profiling, we implicate IRF2 as antagonistic to stemness and show that it binds and regulates active cis-regulatory elements at interferon response and antigen presentation genes. CRISPR-KD of IRF2 in keratinocytes with low stem cell potential increases self-renewal, migration and epidermis formation. These data demonstrate that transcription factor regulatory circuitries, in addition to maintaining cell identity, control plasticity within cell types and offer potential for therapeutic modulation of cell function.

Open in PubMed