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. 2016 Mar 25;44(10):4920–4933. doi: 10.1093/nar/gkw192

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© The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com

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Figure 2. — TREX-2 interaction with ORC. (A) Drosophila embryonic nuclear extract treated with DNase and RNase was immunoprecipitated with affinity purified polyclonal antibodies against TREX-2 or ORC components (indicated on the top) or rabbit Immunoglobulin G (IgG) coupled to protein A-Sepharose beads. Equivalent amounts of the input fraction, and of proteins bound to the immunosorbent (IP) were resolved by SDS-PAGE and analyzed by western blotting with the aforementioned antibodies. Asterisks indicate antibody chains. (B) The recombinant TREX-2 and ORC subunits interact with each other. HA-tagged Xmas-2, PCID2 and ENY2 were co-expressed in S2 cells together with FLAG-tagged ORC subunits, and their interaction was verified in co-immunoprecipitation experiments with antibodies against FLAG or HA epitopes. Asterisk indicates antibody chains. (C) TREX-2 and ORC interact during Drosophila development. Males hemizygous for the e(y)2^u1 mutation of ENY2 and heterozygous for orc3¹ or Orc5² mutations (e(y)2^u1/Y; orc3¹/+ or e(y)2^u1/Y; Orc5²/+) have a decreased viability and an increased frequency of abnormal tergites, comparatively to the control (e(y)2^u1/Y;+/+). Flies homozygous for the P{EP}PCID2^[G18683] mutation and heterozygous for orc3¹ or Orc5² mutations P{EP}PCID2^[G18683]/P{EP}PCID2^[G18683];orc3¹/+ or P{EP}PCID2^[G18683]/P{EP}PCID2^[G18683];Orc5²/+ have decreased viability, comparatively to the control (P{EP}PCID2^[G18683]/P{EP}PCID2^[G18683];+/+).