Research Paper Volume 4, Issue 4 pp 290—304

Figure 4. Phosphorylation of SNEV at S149A is ATM-dependent. (A) ATM inhibition alleviates phosphorylation of SNEV in response to oxidative stress. HDFs were treated with 100 μM H₂O₂ and the specific ATM inhibitor KU-55933. DMSO, the solvent for Ku-99533, was used as negative control. Phosphorylation of SNEV and known ATM target γH2AX was reduced by inhibition of ATM. (B) Phopshorylation of SNEV at S149 is reduced in ATM conditional knockout MEF. Upper panel: wt and ATM -/- MEF were treated with 200 μM H₂O₂ for 1h, harvested, resuspended in 2x SDS loading dye and subjected to SDS PAGE. Western Blot was detected with anti-pSNEV(S149) and anti-βActin to ensure equael loading. Lower panel: Genomic DNA was isolated from wt and ATM lox/lox MEF before and after transfection with Cre and used in a genotyping PCR. The band for deleted ATM is present only after transfection with Cre, nonetheless the deletion is not quantitative and a small portion of ATM is maintained.