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. 2020 May 12;11:2364. doi: 10.1038/s41467-020-16205-9

Fig. 8. ZNF398 is required for somatic cells reprogramming.

Fig. 8

a Experimental strategy for reprogramming by delivery of OSKM (OCT4, SOX2, KLF4, MYC) mRNAs in combination with siRNAs in order to test the requirement of ZNF398 for reprogramming. Scale bars 50 µm. See also “Methods” section for details. b Cell morphology during OSKM reprogramming. Representative images of two independent experiments are shown. See also Supplementary Fig. 8b. c Number of iPSC colonies obtained from 100 cells seeded at day 14 in OSKMNL reprogramming and at day 16 in OSKM reprogramming under the indicated conditions. Bars indicate the mean ± SEM of independent experiments. p-values: unpaired two-tailed Mann–Whitney U test. OSKMNL reprogramming: dots indicate biological replicates (n = 23, 24 and 23 replicates for no siRNA, siCONTROL and siZNF398, respectively) from four independent experiments shown in different shades of colours. OSKM reprogramming: dots indicate biological replicates (n = 15 for replicates scored based on morphology or NANOG signal, n = 8 for replicates scored based on OCT4 signal) from two independent experiments shown in different shades of colours. Source data are provided as a Source Data file. d Immunostaining for OCT4 and NANOG of fibroblasts transfected with OSKM mRNAs and siCONTROL or siZNF398 at day 16. Representative images of two independent experiments are shown. Scale bars 150 µm. e Gene expression analysis by qPCR of hiPSCs (KiPS) (light grey) and fibroblasts (dark grey) serving as controls, and fibroblasts transfected with OSKMNL mRNAs and siCONTROL (light orange) or siZNF398 (dark orange) at day 6 and day 14. Bars indicate the mean of two independent experiments shown as dots. Expression was normalised to the mean of KiPS samples. See also Supplementary Fig. 8a, c. Source data are provided as a Source Data file. f Immunostaining for NANOG and E-Cadherin followed by Phalloidin staining of fibroblasts transfected with OSKM mRNAs and siCONTROL or siZNF398 at day 16. Representative images of two independent experiments are shown. Scale bars 150 µm. g Diagram representing the transcription factors induced by TGF-beta, among which ZNF398 is crucial for the maintenance of pluripotency and the epithelial character of hPSCs.