Defective DNA ligase I in Bloom's syndrome cells. Simultaneous analysis using immunoblotting and the ligase-[32P]AMP adduct assay.
Cells from patients with Bloom's syndrome (BS), an autosomal recessive disorder associated with an increased risk of cancer, exhibit genomic instability. Increased numbers of sister-chromatid exchanges (SCE) and delayed DNA chain maturation are typically observed in BS cells. To elucidate the basis for the previously reported decreased DNA ligase I activity in BS cells, simultaneous immunoblot and activity assays for ligase-[32P]AMP adduct formation were performed on extracts from BS and normal lymphoblastoid cell lines. Immunoblot analysis using antibody to DNA ligase I indicate that the amount of the major reactive protein (98 kDa) in normal and BS cells is similar. However, a 50-90% decrease was observed in the ligase activity of the 98-kDa polypeptide in high-SCE BS cells (HG1514 and GM3403c). In contrast, the activity in low-SCE BS cells (HG1554) did not differ significantly from that in normal cells. The data, together with mixing experiments, indicate that the defect in BS ligase I is due at least in part to the loss of ATP binding and/or hydrolytic activity and not to differences in numbers of protein molecules or inhibitory substances. These results suggest that mutation of the DNA ligase I gene may account for the primary metabolic defect in BS.
Chan
J Y
JY
Department of Molecular Pathology, University of Texas, M. D. AndersonCancer Center, Houston 77030.
Becker
F F
FF
D000249:Adenosine Monophosphate / Q000378:metabolism; D000906:Antibodies; D001816:Bloom Syndrome / Q000201:enzymology*; D002460:Cell Line; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D008970:Molecular Weight; D011118:Polynucleotide Ligases / Q000235:genetics*
D000906:Antibodies; C000607892:LIG1 protein, human; D000249:Adenosine Monophosphate; D011088:DNA Ligases; D011118:Polynucleotide Ligases; D000072481:DNA Ligase ATP