Figure 1
From: MYCL promotes iPSC-like colony formation via MYC Box 0 and 2 domains
MYCL promotes reprogramming more efficiently than c-MYC. (A) Schematic representation of HDF reprogramming with Sendai virus (SeV). HDFs were transduced with SeV carrying KLF4-OCT3/4-SOX2 (KOS), KLF4 (K), and c-MYC or MYCL on day 0. We used an MOI (multiplicity of infection) of 4.3 for each virus. StemFit AK03N without bFGF was used during the transduction and subsequent induction of iPSC-like colonies. We performed immunostaining of the reprogramming HDFs 1 to 7 days after the transduction and analyzed the results using ArrayScan. (B) Representative immunostaining images of reprogramming HDFs stained by anti-TRA-1-60 antibody (green) and Hoechst (blue) 7 days after the transduction. Scale bar, 300 μm. Ph, phase contrast. (C) Proliferation and expression of TRA-1-60 ( +) cells during reprogramming. HDFs were transduced with SeV, including c-MYC or MYCL, and immunostaining was performed from days 1 to 7. The number of total cells was counted as Hoechst-positive cells. Mean ± SD values are shown. n = 3, *p < 0.05 by paired t-test. (D) Schematic representation of HDF reprogramming with episomal plasmid vector (EpiP). HDFs were transduced with EpiP carrying SOX2, KLF4, OCT3/4-shp53, LIN28A, EBNA1, and c-MYC or MYCL. StemFit AK03N without bFGF was used during the transfection and subsequent induction of iPSC-like colonies. We performed flow cytometry of the reprogramming HDFs every three days from 1 to 19 days plus day 21 after the transduction. (E) Representative immunostaining images of reprogramming HDFs stained by anti-TRA-1-60 antibody (green) and Hoechst (blue) 21 days after the transduction. Scale bar, 300 μm. Ph, phase contrast. (F) Proliferation and expression of TRA-1-60 ( +) cells during reprogramming were analyzed by flow cytometry. HDFs were transduced with EpiP, including c-MYC or MYCL. Flow cytometry was performed every three days from days 1 to 19 days plus day 21. Mean ± SD for n = 3, *p < 0.05 and **p < 0.01 by paired t-test. (G) The number of iPSC-like and non-iPSC-like colonies derived from 1 × 105 HDFs transduced with EpiP including c-MYC or MYCL on day 21. Mean ± SD values are shown. n = 3, **p < 0.01 by unpaired t-test. (H) Percentage of CD13 ( +) cells during EpiP reprogramming determined by flow cytometry. Mean ± SD values are shown. n = 3, *p < 0.05 and **p < 0.01 by paired t-test.
