Figure 3.

The level of the H3K56ac protein during the cell cycle. (A) Acid-extracted CCTL12 histones were separated on a short Tris-Tricine gel and Amido Black stained. The locations of the major forms of the core histones are noted. (B) Western blot analysis of H3K56ac at different time points after release from the thymidine block. We did not observe a significant change in the H3K56ac level during the cell cycle in HeLa, HL-60 or CCTL12 cells. In the cancer cell lines, a second band with a lower molecular weight appeared. The band represents the H3 histone after N-terminal degradation. (C) The charts show the level of H3K56ac normalized to the total protein (n=3). The total protein was visualized by Amido Black staining. (D) The charts show the relative ratio of H3K56ac and unacetylated H3K56 in different cell cycle phases obtained by mass spectrometry. H3K56ac was not significantly changed during the cell cycle stages in any of the 3 cell lines. The range of the error bars encompasses the standard deviation values (n = 3). (E) An example of the chromatographic profile of the SRM transitions for the native (YQKSTELLIR) and acetylated (YQK[Ac]STELLIR) forms of the peptide.