Restoration of topoisomerase 2 function by complementation of defective monomers in Drosophila

Amber M Hohl; Morgan Thompson; Alexey A Soshnev; Jianhong Wu; James Morris; Tao-Shih Hsieh; C-Ting Wu; Pamela K Geyer

doi:10.1534/genetics.112.144006

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Restoration of topoisomerase 2 function by complementation of defective monomers in Drosophila

Journal article

Open access

Peer reviewed

Restoration of topoisomerase 2 function by complementation of defective monomers in Drosophila

Amber M Hohl, Morgan Thompson, Alexey A Soshnev, Jianhong Wu, James Morris, Tao-Shih Hsieh, C-Ting Wu and Pamela K Geyer

Genetics (Austin), Vol.192(3), pp.843-856

11/2012

DOI: 10.1534/genetics.112.144006

PMCID: PMC3522162

PMID: 22923380

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Abstract

Type II topoisomerases are essential ATP-dependent homodimeric enzymes required for transcription, replication, and chromosome segregation. These proteins alter DNA topology by generating transient enzyme-linked double-strand breaks for passage of one DNA strand through another. The central role of type II topoisomerases in DNA metabolism has made these enzymes targets for anticancer drugs. Here, we describe a genetic screen that generated novel alleles of Drosophila Topoisomerase 2 (Top2). Fifteen alleles were obtained, resulting from nonsense and missense mutations. Among these, 14 demonstrated recessive lethality, with one displaying temperature-sensitive lethality. Several newly generated missense alleles carry amino acid substitutions in conserved residues within the ATPase, Topoisomerase/Primase, and Winged helix domains, including four that encode proteins with alterations in residues associated with resistance to cancer chemotherapeutics. Animals lacking zygotic Top2 function can survive to pupation and display reduced cell division and altered polytene chromosome structure. Inter se crosses between six strains carrying Top2 missense alleles generated morphologically normal trans-heterozygous adults, which showed delayed development and were female sterile. Complementation occurred between alleles encoding Top2 proteins with amino acid substitutions in the same functional domain and between alleles encoding proteins with substitutions in different functional domains. Two complementing alleles encode proteins with amino acid substitutions associated with drug resistance. These observations suggest that dimerization of mutant Top2 monomers can restore enzymatic function. Our studies establish the first series of Top2 alleles in a multicellular organism. Future analyses of these alleles will enhance our knowledge about the contributions made by type II topoisomerases to development.

Mutagenesis

Mutation

Phenotype

DNA Topoisomerases, Type II - metabolism

Polytene Chromosomes

Male

Fertility - genetics

Animals

Protein Interaction Domains and Motifs - genetics

Alleles

DNA Topoisomerases, Type II - chemistry

DNA Topoisomerases, Type II - genetics

Female

Drosophila - metabolism

Gene Order

Amino Acid Substitution

Drosophila - genetics

Details

Title: Subtitle: Restoration of topoisomerase 2 function by complementation of defective monomers in Drosophila
Creators: Amber M Hohl - Graduate Program in Genetics, University of Iowa, Iowa City, Iowa 52242, USA
Morgan Thompson
Alexey A Soshnev
Jianhong Wu
James Morris
Tao-Shih Hsieh
C-Ting Wu
Pamela K Geyer
Resource Type: Journal article
Publication Details: Genetics (Austin), Vol.192(3), pp.843-856
Publisher: United States
DOI: 10.1534/genetics.112.144006
PMID: 22923380
PMCID: PMC3522162
ISSN: 0016-6731
eISSN: 1943-2631
Grant note: R01 GM085169 / NIGMS NIH HHS R01 GM042539 / NIGMS NIH HHS GM085169 / NIGMS NIH HHS GM42539 / NIGMS NIH HHS
Language: English
Date published: 11/2012
Academic Unit: Obstetrics and Gynecology; Biochemistry and Molecular Biology
Record Identifier: 9984025260902771

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