Figure 5.
Figure 5. Requirement of LIG4 for DSB-stimulated DNA-PKcs autophosphorylation in cells and contribution of Cer-XLF. (A and B) Nalm6 control and N114P2 LIG4-null pre-B cells were treated for 1 h with increasing doses of Calicheamicin γ1 (Cali) as indicated (A) or with 160 pM Cali followed by incubation in fresh medium under normal growth conditions for the specified time (B). Whole-cell extracts were heat denatured, separated on SDS-PAGE gels, and electrotransferred onto membranes that were blotted with antibodies as indicated. Protein samples were separated on 8% acrylamide SDS-PAGE gels for standard separation or 15% for γ-H2AX isolation. (C) Western blotting on the whole-cell extracts of BuC control and BuS Cer-XLF- mutant fibroblasts after treatment with increasing doses of Cali as indicated for 1 h. NT, not treated; PhS59, phosphorylation of SAF-A on S59.

Requirement of LIG4 for DSB-stimulated DNA-PKcs autophosphorylation in cells and contribution of Cer-XLF. (A and B) Nalm6 control and N114P2 LIG4-null pre-B cells were treated for 1 h with increasing doses of Calicheamicin γ1 (Cali) as indicated (A) or with 160 pM Cali followed by incubation in fresh medium under normal growth conditions for the specified time (B). Whole-cell extracts were heat denatured, separated on SDS-PAGE gels, and electrotransferred onto membranes that were blotted with antibodies as indicated. Protein samples were separated on 8% acrylamide SDS-PAGE gels for standard separation or 15% for γ-H2AX isolation. (C) Western blotting on the whole-cell extracts of BuC control and BuS Cer-XLF- mutant fibroblasts after treatment with increasing doses of Cali as indicated for 1 h. NT, not treated; PhS59, phosphorylation of SAF-A on S59.

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