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. 2018 Jan 5;25(3):600–615. doi: 10.1038/s41418-017-0002-6

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© ADMC Associazione Differenziamento e Morte Cellulare 2018

Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License, which permits any non-commercial use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. If you remix, transform, or build upon this article or a part thereof, you must distribute your contributions under the same license as the original. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-sa/4.0/.

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Fig. 7 — HDAC4 decreases Notch1 activity in the presence of SUMO. a Sumoylated Notch1 binds to CSL and remains bound to DNA. n = 4. b Blocking the expression of HDAC4 by siRNA increases Notch1 activity. Notch1 activity was measured from HeLa cells by the 12×CSL luciferase reporter reflecting the activity of the Notch1 signaling pathway. The values shown are normalized luciferase units. The difference in Notch1 activity is greater in samples with overexpressed SUMO1 (S1) n = 3. c HDAC4 represses the activity of wild-type GFP-Flag-NICD1 (WT) more than it represses the activity of the K1774/1780/1781/1782R GFP-Flag-NICD1 (TSM). n = 3. d SUMO increases Notch1–HDAC4 interaction. Control samples without antibody during IP are indicated with an “M” referring to mock. n = 3. e Consistent with the results in (c), K1774/1780/1781/1782R GFP-Flag-NICD1 (NICD TSM) decreases NICD1–HDAC4 interaction compared to wildtype NICD1 (NICD WT). f Notch interacts with HDAC4 on the DNA. n = 3. g The levels of HDAC4 as related to Notch in the immunoprecipitates in (e) were quantified. Values indicate the average of three independent experiments. n = 3. h–j Inhibition of HDAC4 expression by siRNA increases, and HDAC4 overexpression decreases the expression of Notch1 (N1) target genes Hes1, Hey1, and Hey2 in the presence of SUMO1 (S1). The relative expression of mRNA was analyzed by quantitative RT-PCR. n = 3