SCAN1 mutant Tdp1 accumulates the enzyme--DNA intermediate and causes camptothecin hypersensitivity.
Tyrosyl-DNA phosphodiesterase (Tdp1) catalyzes the hydrolysis of the tyrosyl-3' phosphate linkage found in topoisomerase I-DNA covalent complexes. The inherited disorder, spinocerebellar ataxia with axonal neuropathy (SCAN1), is caused by a H493R mutation in Tdp1. Contrary to earlier proposals that this disease results from a loss-of-function mutation, we show here that this mutation reduces enzyme activity approximately 25-fold and importantly causes the accumulation of the Tdp1-DNA covalent reaction intermediate. Thus, the attempted repair of topoisomerase I-DNA complexes by Tdp1 unexpectedly generates a new protein-DNA complex with an apparent half-life of approximately 13 min that, in addition to the unrepaired topoisomerase I-DNA complex, may interfere with transcription and replication in human cells and contribute to the SCAN1 phenotype. The analysis of Tdp1 mutant cell lines derived from SCAN1 patients reveals that they are hypersensitive to the topoisomerase I-specific anticancer drug camptothecin (CPT), implicating Tdp1 in the repair of CPT-induced topoisomerase I damage in human cells. This finding suggests that inhibitors of Tdp1 could act synergistically with CPT in anticancer therapy.
Interthal
Heidrun
H
Department of Microbiology, School of Medicine, University of Washington, Seattle, WA 98195-7242, USA.
Chen
Hong Jing
HJ
Kehl-Fie
Thomas E
TE
Zotzmann
Jörg
J
Leppard
John B
JB
Champoux
James J
JJ
D019943:Amino Acid Substitution; D002166:Camptothecin / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005434:Flow Cytometry; D006801:Humans; D009154:Mutation; D009369:Neoplasms / Q000188:drug therapy; D010727:Phosphoric Diester Hydrolases / Q000235:genetics* / Q000378:metabolism; D016196:S Phase / Q000187:drug effects
D004791:Enzyme Inhibitors; D010727:Phosphoric Diester Hydrolases; C498508:TDP1 protein, human; C122147:tyrosyl-DNA phosphodiesterase; D002166:Camptothecin