Fig. 4 | Communications Biology

Fig. 4

From: Sumoylation regulates the stability and nuclease activity of Saccharomyces cerevisiae Dna2

Fig. 4

Sumoylation targets Dna2 for degradation. a A representative western blot comparing the levels of Dna2 vs. non-sumoylatable Dna26K proteins. Cells (JKM139 background) were synchronized at G1, and released for indicated times. Ponceau-stained membrane is a control for equal loading. Protein levels from asynchronous cells (Async) are also shown. b Quantitation of Dna2 or Dna26K protein levels from the experiments such as shown in a. The values are expressed relative to Dna26K levels at S-G2. Averages shown, n = 3; whiskers, s.e.m. c Real-time PCR was used to obtain cycle threshold levels (relative to Actin, ΔCt) to quantitate DNA2 and RAD27 mRNA levels from DNA2 and dna26K cells (JKM139 background). Averages shown, n = 3; whiskers, s.e.m. d Comparison of Dna2 and Dna26K levels in whole cell (20 μg) and nuclear extracts (3.8 μg) (JKM139 background) by western blotting. Asterisk indicates a putative Dna2 truncation product. e Quantitative analysis of Dna2-YFP or Dna26K-YFP foci. Cells (yLK354 and yLK388, W303 background) were treated with methyl-methanesulfonate (MMS, 0.015% for 3 h) and subjected to microscopy analysis. The fraction of cells showing Yellow Fluorescent Protein foci is shown. Average and SEM of three independent experiments are shown (n > 160 cells per strain). Statistical significance relative to the wild type was determined by Fisher’s exact test from the total numbers of cells with or without foci from all experiments

Back to article page