Role for Nucleolin/Nsr1 in the Cellular Localization of Topoisomerase I

Troy K Edwards,Ahamed Saleem,Jeffrey A Shaman,Tracey Dennis,Claudia Gerigk,Elder Oliveros,Marc R Gartenberg,Eric H Rubin

doi:10.1074/jbc.m006628200

Troy K Edwards, Ahamed Saleem + Show 6 more

Open Access

https://doi.org/10.1074/jbc.m006628200

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Abstract

Nucleolin functions in ribosome biogenesis and contains an acidic N terminus that binds nuclear localization sequences. In previous work we showed that human nucleolin associates with the N-terminal region of human topoisomerase I (Top1). We have now mapped the topoisomerase I interaction domain of nucleolin to the N-terminal 225 amino acids. We also show that the Saccharomyces cerevisiae nucleolin ortholog, Nsr1p, physically interacts with yeast topoisomerase I, yTop1p. Studies of isogenic NSR1(+) and Deltansr1 strains indicate that NSR1 is important in determining the cellular localization of yTop1p. Moreover, deletion of NSR1 reduces sensitivity to camptothecin, an antineoplastic topoisomerase I inhibitor. By contrast, Deltansr1 cells are hypersensitive to the topoisomerase II-targeting drug amsacrine. These findings indicate that nucleolin/Nsr1 is involved in the cellular localization of Top1 and that this localization may be important in determining sensitivity to drugs that target topoisomerases.

Highlights

Human Top11 is a nuclear protein involved in the regulation of DNA structure and is the target of an important new class of antineoplastic drugs, the camptothecins [1, 2]
The N Terminus of Nucleolin Is Necessary and Sufficient for Top1 Binding—To identify regions of nucleolin involved in Top1 binding, we generated a recombinant protein consisting of nucleolin fused to the C terminus of GST
We considered the possibility that nucleolin functions to recruit Top1 to sites of rDNA transcription

Summary

Introduction

Human Top is a nuclear protein involved in the regulation of DNA structure and is the target of an important new class of antineoplastic drugs, the camptothecins [1, 2]. Current models invoke interactions between ternary complexes and replication or transcriptional machinery as being important in conversion of ternary complexes to lethal forms of DNA damage [12, 13] It is not known whether physical interactions between Top and other proteins (including proteins involved in replication or transcription) are important in the cytotoxic effects of CPT. Top rapidly redistributes from the nucleolus to the nucleus or cytoplasm and is degraded after cellular exposure to CPT [15,16,17,18,19] These alterations may confer transient cellular resistance to CPT and could be mediated by interactions between Top and other proteins. Our results indicate that NSR1 is involved in the cellular localization of yTop1p and that loss of NSR1 alters cellular sensitivity to topoisomerase-targeting drugs but not to DNA damaging agents in general

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