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. 2017 Aug 15;31(16):1693–1703. doi: 10.1101/gad.302000.117

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© 2017 Maezawa et al.; Published by Cold Spring Harbor Laboratory Press

This article is distributed exclusively by Cold Spring Harbor Laboratory Press for the first six months after the full-issue publication date (see http://genesdev.cshlp.org/site/misc/terms.xhtml). After six months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.

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Figure 6. — RNF2 interacts with SALL4, and RNF2 and SALL4 work together for gene activation. (A) Coimmunoprecipitation of RNF2 and SALL4 using wild-type testes at 14–19 d of age. (B) Dual-luciferase reporter assays using mK4 cells. Regulatory regions near TSSs of the Sall4, Mael, and Tdrkh genes were examined. (miniP) Minimal promoter. The relative luciferase activities (Nluc/Fluc) were normalized to Nluc/Fluc ratios from mock experiments in mK4 cells. (C,D) Dual-luciferase reporter assays using embryonic stem (ES) cells. Leukemia inhibitory factor (LIF) was used to maintain undifferentiated ES cells, and differentiation was induced through the withdrawal of LIF and the addition of vitamin C (VitC) and/or retinoic acid (RA). Relative luciferase activities (Nluc/Fluc) were normalized to Nluc/Fluc ratios from mock experiments in undifferentiated ES cells. (*) P < 0.05; (**) P < 0.005; (***) P < 0.001, unpaired t-test. Data are represented as mean ± SEM. Three to five independent experiments were performed. (E) Model of action of RNF2 and SALL4 for activation of germline genes, in which RNF2 activates transcription of Sall4, and RNF2 and SALL4 together occupy target genes for activation.