Fig. 5: STUbL and Cullin8 protect replication-associated recombination structures by preventing unscheduled Mms4/Mus81-dependent processing.

a 2D gel profiles of recombination intermediates isolated from cells of the indicated genotype. Cells were synchronized in G1 phase with α-factor and then released in medium containing 0.033% MMS. Tetracycline 1 mM was added during G1 arrest and release. Samples for 2D gel analysis were collected at the indicated timepoints and levels of HA-tagged Sgs1 were monitored by western blot. Immunodetection of Pgk1 served as a loading control. Cell cycle progression of the cells during the experiment was followed by flow cytometric analysis. 1N and 2N in gray indicate G1 and G2/M phases, respectively. Replication intermediates were digested with NcoI and visualized using a radioactively labeled probe specific for ARS305 in 2D gel electrophoresis. Red arrows indicate X-shaped replication intermediates, the relative enrichment of which is shown in the quantified plots. Signal intensities were normalized to the monomer spot and shown relative to the highest value assigned as 100%. b 2D gel profiles of recombination intermediates isolated from cells of the indicated genotype. Experimental set-up as in a. The replication intermediates were digested with EcoRV and HindIII and visualized using a radioactively labeled probe specific for ARS301. Source data are provided as a Source data file.