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. 2013 Nov 14;42(3):1628–1643. doi: 10.1093/nar/gkt1041

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© The Author(s) 2013. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 4. — The level of trimethylated UBF (Tri-Met UBF) is regulated by ESET. (A) The immunoreactivity of Myc-tagged ESET and H3K9me3 was increased by doxycycline (Doxy). (B) The protein level of Myc-tagged ESET was robustly induced by Doxy (n = 3). (C) The increased levels of ESET and TMH3K9 were found in nuclear and nucleolar fractions in response to Doxy. The purity of subcellular fraction was determined by specific antibody as follows: Lamin B, a nucleus marker; Fibrillarin, a nucleolus marker. (D) The induction of ESET by doxycycline for the indicated period of time increased the expression trimethylated UBF in a time-dependent manner. (E) Cells were treated with doxycycline for 36 h (ESET ON) and then cells were washed of doxycycline and switched to the normal fresh media for 36 h (ESET OFF). The level of Tri-Met UBF was increased after 36 h of the ESET ON condition and was decreased to the basal levels after 36 h of the ESET OFF condition and vice versa. This change was depended on the induced or reduced of ESET. The whole blot of Tri-Met UBF in (E) is presented in the Supplementary Figure S8. (F) H3K9me3 immunoreactivity was markedly induced in ESET ON cells compared with ESET OFF cells. Scale bar: 10 µm.