hela s3 tet off cells  (TaKaRa)

Journal:

Article Title: Structural and Functional Conservation of the NuA4 Histone Acetyltransferase Complex from Yeast to Humans

doi: 10.1128/MCB.24.5.1884-1896.2004

Figure Lengend Snippet: SANT domain-containing protein DMAP1 is a stable subunit of the human NuA4 complex. (A) Amino acid sequence alignment between yeast NuA4 subunit Eaf2, human DMAP1, and a Drosophila homologous protein (dEaf2, accession number AAF57436). The conserved SANT domain region is indicated. (B) DMAP1 associates with the same set of proteins as Tip60. Nuclear extracts from FLAG-Tip60-TAP, FLAG-DMAP1-TAP, or mock-transduced HeLa S3 cells were partially purified over IgG-Sepharose beads, and the TEV eluate was analyzed by Western blotting as in Fig. ​Fig.1B.1B. (C) Triple affinity purification of DMAP1 identifies distinct non-HAT complex(es). The DMAP1-containing complexes were purified as in Fig. ​Fig.22 and analyzed on a gel by quantitative Sypro Ruby staining. Proteins identified by Western analysis or tandem mass spectrometry are labeled on the right. New proteins not identified in the Tip60 purification are in italics. The positions of MRG15 and Tip60 based on Western analysis of the previous fraction are given in parentheses. The asterisk indicates a nonspecific band. A significant amount of GAS41 protein was also detected but was run out of the gel presented here (see Western signal in panel B). The numbers of specific peptide sequences obtained by tandem mass spectrometry analysis performed as described for Fig. ​Fig.22 are as follows: 6 for TRRAP, 8 for p400/hDomino, 7 for SRCAP (2,971 aa), 4 for Brd8, 13 for DMAP1, 4 for YL-1 (364 aa), 10 for RUVBL1, 14 for RUVBL2, 5 for BAF53a, and 5 for actin.

Article Snippet: MCF7 and HeLa S3 tet-off cell lines were purchased from Clontech and cultured in Dulbecco modified Eagle medium supplemented with 10% fetal calf serum.

Techniques: Sequencing, Purification, Western Blot, Affinity Purification, Staining, Mass Spectrometry, Labeling