Abstract
DNA topoisomerase I (topo I) is involved in the regulation of DNA supercoiling, gene transcription, recombination, and DNA repair. The anticancer agent camptothecin specifically targets topo I. The mechanisms responsible for the regulation of topo I in cells, however, are not known. This study demonstrates that c-Abl-dependent phosphorylation up-regulates topo I activity. The c-Abl SH3 domain bound directly to the N-terminal region of topo I. The results demonstrate that c-Abl phosphorylated topo I at Tyr268 in core subdomain II. c-Abl-mediated phosphorylation of topo I Tyr268 in vitro and in cells conferred activation of the topo I isomerase function. Moreover, activation of c-Abl by treatment of cells with ionizing radiation was associated with c-Abl-dependent phosphorylation of topo I and induction of topo I activity. The functional significance of the c-Abl/topo I interaction is supported by the findings that (i) mutant topo I(Y268F) exhibited loss of c-Abl-induced topo I activity, and (ii) c-Abl-/- cells were deficient in the accumulation of protein-linked DNA breaks. In addition, loss of topo I phosphorylation in c-Abl-deficient cells conferred resistance to camptothecin-induced apoptosis. These findings collectively support a model in which c-Abl-mediated phosphorylation of topo I is functionally important to topo I activity and sensitivity to topo I poisons.
Highlights
Eukaryotic type I topoisomerases function in the relaxation of negatively and positively supercoiled DNA [1,2,3,4]
Because c-Abl is activated by DNA strand breaks [45] and Topoisomerase I (topo I) is involved in the generation of these lesions [35], we investigated whether c-Abl interacts with topo I
Immunoprecipitation of the lysates with antiphospho-Tyr antibody and immunoblot analysis of the supernatants demonstrated a 25% decrease in the amount of topo I. These findings indicate that ϳ1 of 4 topo I molecules is subject to tyrosine phosphorylation and/or that topo I coprecipitates with other tyrosine-phosphorylated proteins
Summary
Vol 279, No 50, Issue of December 10, pp. 51851–51861, 2004 Printed in U.S.A. Phosphorylation of DNA Topoisomerase I by the c-Abl Tyrosine Kinase Confers Camptothecin Sensitivity*. Loss of topo I phosphorylation in c-Abl-deficient cells conferred resistance to camptothecin-induced apoptosis These findings collectively support a model in which c-Abl-mediated phosphorylation of topo I is functionally important to topo I activity and sensitivity to topo I poisons. The DNA-dependent protein kinase catalytic subunit and ATM are members of a family of phosphatidylinositol 3-kinase-like enzymes involved in the regulation of the cell cycle, recombination, control of telomere length, and the DNA damage response [44]. Other studies have demonstrated that nuclear c-Abl interacts with the human catalytic subunit of telomerase, TERT, and thereby regulates telomere length [52] These findings have supported a role for nuclear c-Abl in converting DNA damage into signals that control the genotoxic stress response. We show that c-Abl-mediated activation of topo I is important, at least in part, in imparting cellular sensitivity to CPT
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
