BCR-ABL–positive CML cell lines show down-regulation of major NHEJ proteins, Artemis, and DNA ligase IV, and up-regulation of alternative NHEJ proteins, DNA ligase IIIα, and WRN. (Ai) Western blotting using nuclear extracts from 2 BCR-ABL–positive CML cell lines (K562 and MEG01), and 3 EBV-transformed B-cell lines, established from healthy individuals (NC3, NC10, and NC108), and (Aii) 1 BCR-ABL–negative CML cell line (MO7e). Artemis shows down-regulation (4- to 7-fold) in BCR-ABL–positive CML cell lines. Either actin or Ku86 was used as loading control. (B) Western blotting using nuclear extracts from K562 shows 2- to 3-fold decrease in DNA ligase IV expression, compared with EBV-transformed B-cell lines (NC3 and NC10). Ku86 was used as loading control. (C) Western blotting using nuclear extracts from 2 BCR-ABL–positive CML cell lines (K562 and MEG01) and 3-EBV transformed B-cell lines (NC3, NC10, and NC108). CML cell lines show up-regulation of DNA ligase IIIα (3- to 6-fold). PARP1 expression is unchanged. Werner syndrome protein, WRN, shows up-regulation (4- to 7-fold) in CML cell lines. Actin was used as loading control. (D) BCR-ABL–negative MO7e shows down-regulation of DNA ligase IIIα and WRN compared with BCR-ABL–positive P210MO7e and K562. Western blotting using nuclear extracts from 2 BCR-ABL–positive cell lines (K562 and P210MO7e) and BCR-ABL–negative cell lines (U937 and MO7e). Actin was used as loading control. (E) Primary CML bone marrow mononuclear cells show down-regulation of DNA ligase IIIα and WRN. Western blotting using nuclear extracts from CML cell line (K562) and 2 CML patients (CML1, CML2) at diagnosis. Patient CML1 shows decreased DNA ligase IIIα and WRN expression with decreased levels of BCR-ABL positivity by FISH. Patient CML2 shows increased expression of DNA ligase IIIα and WRN, with 100% BCR-ABL positivity. BCR-ABL–positive CML cell line K562 was used as a positive control. Ku86 and actin were used as loading controls.