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. 2017 Aug 30;7:9932. doi: 10.1038/s41598-017-09494-6

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits 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. 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/4.0/.

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Consequences of acute E2f3 induction in adult tissues. (a) Quantitative RT-PCR analysis of E2f1, Cdc2 and CyclinA in adult control (n = 5 (spleen, lung), n = 4 (liver)) and E2f3 ^TRE/TRE; Rosa26 ^rtTA/rtTA (n = 3) mouse tissues following administration of doxycycline for three days. Expression is normalized to HPRT and relative to the mean of the control samples within each tissue. Error bars, s.e.m. Two-tailed t-test; control vs rtTA/rtTA: **P = 0.01 (CyclinA, liver), 0.05* (Cdc2, liver), 0.05* (E2f1, spleen). (b) Immunohistochemical staining of phosphorylated histone 3 (p-H3) in the spleen and Ki67 in the liver and lung of control and E2f3 ^TRE/TRE; Rosa26 ^rtTA/rtTA mice following administration of doxycycline for three days. Scale bar 100 µm. (c) Quantification of p-H3 positive nuclei per field of view of the spleen (red pulp) or Ki67 positive nuclei per field of view in the liver (hepatocytes) and lung isolated from control (n = 5(spleen, lung), n = 9(liver)), E2f3 ^TRE/TRE; Rosa26 ^rtTA/+ (n = 3(spleen), n = 4(liver, lung)) and E2f3 ^TRE/TRE; Rosa26 ^rtTA/rtTA (n = 4(spleen, lung), n = 3(liver)) mice following administration of doxycycline for three days. Mean of 5 images per mouse; error bars, s.e.m. Mann-Whitney test; control vs. rtTA/+, *P = 0.03 (liver); control vs. rtTA/rtTA, ***P = 0.005 (liver), 0.03* (lung). (d) Quantification of Ki67 positive nuclei per field of view in liver (hepatocytes) isolated from control (n = 9) and E2f3 ^TRE/TRE; Rosa26 ^rtTA/rtTA mice after 3 (n = 3) or 60 (n = 5) days continuous doxycycline administration. Mean of 5 images per mouse; error bars, s.e.m. Control and rtTA/rtTA quantifications from panel C are included for comparison. Mann-Whitney test; control vs. 60days, **P = 0.005. Controls for (a–d) include E2f3^TRE/TRE on and off doxycycline and E2f3 ^TRE/TRE; Rosa26 ^rtTA/+ and E2f3 ^TRE/TRE; Rosa26 ^rtTA/rtTA maintained in the absence of doxycycline.