Figure 5
From: Baf53a is involved in survival of mouse ES cells, which can be compensated by Baf53b
Transfection of wild-type Baf53a (WT), but not mutant Baf53a (M3), rescues the viability of Baf53a-deficient ES cells. (A) Schematic view of Baf53a M3 mutant. Three amino acid residues (glutamic acid (E), arginine (R), and arginine (R)) of wild-type Baf53a were changed into three alanine (A) residues. (B) Morphological signatures of Baf53a-expressing Baf53a cKO ES cells. Baf53a cKO ES cells were transfected with expression vectors encoding empty (ev), Baf53a WT, or Baf53a M3. Each transfected ES cells were divided into two dishes at 48 h after transfection and cultured with or without 1 μg/mL Tet for 4 days and their cell morphologies were observed. Scale bar is 250 μm. (C,D) Colony formation assay. (C)Baf53a WT- or Baf53a M3-expressing Baf53a cKO ES cells and control cells described in panel B were cultured for 6 days with or without 1 μg/mL Tet; and these samples were stained by crystal violet. (D) These colonies were dissolved in 1.0% SDS solution and absorbance was measured at 600 nm to quantify the number of surviving colonies. The representative results shown are expressed as the means ± standard deviation of three independent experiments. (E) Expression of exogenous Myc-tagged Baf53a protein. Expression of either Myc-Baf53a WT or Myc-Baf53a M3 was confirmed by Western blot at 2 days after Tet treatment. αTubulin was used as a loading control. The result is representative of three independent experiments.
