Fig. 6: SIRT3 regulates histone Kbhb levels in a class-selective manner in nucleus.
From: Molecular basis for hierarchical histone de-β-hydroxybutyrylation by SIRT3
a Fluorescence analysis of SIRT3 in HEK293 cells. Scale bars, 5 μm. b A HEK293 cell was analyzed with DAPI (blue) and SIRT3-EGFP (green) and scanned at 0.38 μm increments along the z axis using confocal microscopy (top image). The fluorescence intensities midlane in the cell in the XY axis are shown (middle and bottom images). c Schematic diagram of subcellular fractionation procedures used in this assay (Thermo Scientific, CAT# 78840: Subcellular Protein Fractionation Kit for Cultured Cells). Note that all fractions are adjusted to the same volume before analysis to allow direct abundance comparison. d, e Western blots of HEK293 cell extracted fractions from wild-type cells (d) or after transfection with SIRT3-EGFP (e). CE, cytoplasmic extracts; ME, membrane extracts; NE, soluble nuclear extracts; CB, chromatin-bound extracts; PE, cytoskeletal extracts. f Immunoblotting results of full-length and short form SIRT3 in nuclear (NE) and none nuclear (Non. NE) fractions. g Immunoblotting results of SIRT3 levels after bhbNa treatment in chromatin-bound (CB) fractions. h Immunoblot of histone Kbhb levels of annotated marks in SIRT3 overexpressed HEK293 cells
