112 | Chapter 4 Supplemental Figure 4. GADD45a expression is not affected by DNA damage and inhibition of ATM kinase activity. (A) Western blot analysis of STI571-treated (72h) AH2 (Artemis-/-) and R2K.ART (Artemis-/- / Rag2-/-) mouse v-Abl cell lines. DNA damage (2h treatment with 50 ng/mL NCS) did not affect GADD45a protein levels. Effective induction of DNA damage was shown by phosphorylation of ATM (Ser1981), which was inhibited by treatment of cells with the ATM kinase inhibitor KU55933 2h prior to induction of DNA damage by NCS. The b-actin blot was used to control for loading. (B) Western blot analysis of STI571-treated (72h) BV173 pre-B cells. DNA damage did not affect GADD45a protein levels. The β-actin blot was used to control for loading. (C) Realtime quantitative RT-PCR analysis of Gadd45a mRNA expression in STI571-treated (72h) AH2 and R2K.ART mouse v-Abl cell lines. Gadd45a mRNA expression was not affected by DNA damage and inhibition of ATM kinase activity. (n=3, independent replicate experiments). (D) Realtime quantitative RT-PCR analysis of GADD45a mRNA expression in BV173 pre-B cells. DNA damage did not affect GADD45a mRNA expression levels. (n=4, independent replicate experiments).
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