Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2013 Oct 15;27(20):2259–2273. doi: 10.1101/gad.223180.113

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© 2013 Ragland 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 3.0 Unported), as described at http://creativecommons.org/licenses/by-nc/3.0/.

PMC Copyright notice

Figure 6. — RNF4 and PLK1 function in processive replication after restart. (A) Replication restart at differing time points after APH removal. Short pulses of EdU from 0 to 30, 30 to 60, and 150 to 180 min after APH removal were applied to cells to quantify the frequency of replication recovery by flow cytometry. Note that the majority of replication recovery in RNF4-suppressed Atr^Δ/− cells occurs within the first hour after APH removal, which is the total length of replication recovery assayed in all other experiments described (D–F and Figs. 1, 3, 4). (B) Representative examples of pre-APH BrdU (red) and post-APH EdU (green) labeling of combed DNA in Atr^flox/− cells and Atr^Δ/− cells with RNF4 suppression. Cells were BrdU-labeled for 15 min, treated with 5 μM APH for 6 h, and then EdU-labeled for 20 min immediately after APH removal. (C) Quantification of EdU-labeled replication tracks 20, 30, and 45 min after APH removal in Atr^flox/− cells and Atr^Δ/− cells with RNF4 suppression. The experiment was conducted as described in B, with the 20-min time point representing the earliest replication tracks after APH removal. Note that track lengths after APH removal in RNF4-suppressed Atr^Δ/− cells fail to extend further than that observed in the initial 20 min of labeling. (D) Quantification of replication restart by DNA combing in cells subjected to combined suppression of RNF4 and PLK1. ATR deletion, RNF4 suppression, PLK1 inhibition, and subsequent DNA combing were performed and scored as described in Figures 3 and 4. (E) Replication recovery on a population basis in cells subjected to combined or separate suppression of RNF4 and PLK1. APH treatment was for 6 h before removal and subsequent EdU labeling; replication recovery was quantified by flow cytometry. Note that although suppression of RNF4 or PLK1 singly permits replication recovery, suppression of both together does not. (F) Quantification of replication restart shown in E.