Fig 2: Importins KPNA2, TNPO1 and IPO7 mediated basal nuclear import of SIRT2. (A) Images of HeLa cells stably expressing SIRT-GFP (green) and transfected with stated siRNAs were left untreated or treated with LMB (20 nM) for 1 hour. Nuclei (blue) were stained using DAPI. Scale bar, 10 µm. (B) Graphs show nuclear: whole cell intensity ratio of SIRT2-GFP. Each data point represents a single cell measured from 3 independent experiments. Statistical significance was determined between importin siRNA treated (blue) and scrambled siRNA cells (red) treated with LMB as determined by one-way ANOVA with Dunnet correction for multiple comparisons ***p < 0.0001.

Fig 3: The C-terminus of SIRT2 negatively regulates importin binding and nuclear accumulation. (A) Schematics of full-length SIRT2 (SIRT2FL) and SIRT2 lacking its unstructured C-terminus (SIRT21–356). GFP, SIRT2FL or truncated SIRT21–356 were immunoprecipitated for 1 hr. Cell lysates (INPUT) and IP fractions were immunoblotted for with antibodies against GFP and (B) KPNA2, (C) IPO7 and TNPO1 (D) CBP which served as control. Data are representative of 3 experiments. Uncropped blots are presented in Supplementary S3. (E) Quantification of live-cell imaging of HeLa cells expressing SIRT2FL (green) or SIRT21–356 (orange) treated with LMB (20 nM). Graph shows the mean fluorescence intensity in the nucleus over time from, images were acquired every 5 mins. Error bars represent SEM of 4 independent experiments. Lines represents the slope of a regression line for each data set. (F) Representative still images used for analysis in (E). Scale bar, 10 µm.