Am J Cancer Res 2013;3(5):478-489
Original Article
Functional analysis of Drosophila DNA polymerase ε p58 subunit
Ritsuko Sahashi, Risa Matsuda, Osamu Suyari, Mieko Kawai, Hideki Yoshida, Sue Cotterill, Masamitsu Yamaguchi
Department of Applied Biology, Insect Biomedical Research Center, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585,
Japan; Department Basic Medical Sciences, St Georges University London, Cranmer Terrace, London SW17 0RE, UK; Current
address: Environmental Research Laboratory of Public Health, Kankyo Eisei Yakuhin Co. Ltd., 3-6-2 Hikaridai, Seika-cho,
Soraku-gun, Kyoto 619-0237, Japan. Equal contributors.
Received September 26, 2013; Accepted October 20, 2013; Epub November 1, 2013; Published November 15, 2013
Abstract: DNA polymerase ε (polε) plays a central role in DNA replication in eukaryotic cells, and has been suggested to the
main synthetic polymerase on the leading strand. It is a hetero-tetrameric enzyme, comprising a large catalytic subunit (the A
subunit ~250 kDa), a B subunit of ~60 kDa in most species (~80 kDa in budding yeast) and two smaller subunits (each ~20
kDa). In Drosophila, two subunits of polε (dpolε) have been identified. One is the 255 kDa catalytic subunit (dpolεp255), and the
other is the 58 kDa subunit (dpolεp58). The functions of the B subunit have been mainly studied in budding yeast and
mammalian cell culture, few studies have been performed in the context of an intact multicellular organism and therefore its
functions in this context remain poorly understood. To address this we examined the in vivo role of dpolεp58 in Drosophila. A
homozygous dpolεp58 mutant is pupal lethal, and the imaginal discs are less developed in the third instar larvae. In the eye
discs of this mutant S phases, as measured by BrdU incorporation assays, were significantly reduced. In addition staining with
an anti-phospho histone H3 (PH3) antibody, (a marker of M phase), was increased in the posterior region of eye discs, where
usually cells stop replicating and start differentiation. These results indicate that dpolεp58 is essential for Drosophila
development and plays an important role in progression of S phase in mitotic cell cycles. We also observed that the size of
nuclei in salivary gland cells were decreased in dpolεp58 mutant, indicating that dpolεp58 also plays a role in endoreplication.
Furthermore we detect a putative functional interaction between dpolε and ORC2 in discs suggesting that polε plays a role in the
initiation of DNA replication in Drosophila. (ajcr0000232).
Keywords: DNA polymerase ε B subunit, Drosophila melanogaster
Address correspondence to: Masamitsu Yamaguchi, Department of Applied Biology, Kyoto Institute of Technology,
Matsugasaki, Sakyo-ku, Kyoto 606-8585, Japan. Tel: +81 75 724 7781; E-mail: myamaguc@kit.ac.jp
Original Article
Functional analysis of Drosophila DNA polymerase ε p58 subunit
Ritsuko Sahashi, Risa Matsuda, Osamu Suyari, Mieko Kawai, Hideki Yoshida, Sue Cotterill, Masamitsu Yamaguchi
Department of Applied Biology, Insect Biomedical Research Center, Kyoto Institute of Technology, Sakyo-ku, Kyoto 606-8585,
Japan; Department Basic Medical Sciences, St Georges University London, Cranmer Terrace, London SW17 0RE, UK; Current
address: Environmental Research Laboratory of Public Health, Kankyo Eisei Yakuhin Co. Ltd., 3-6-2 Hikaridai, Seika-cho,
Soraku-gun, Kyoto 619-0237, Japan. Equal contributors.
Received September 26, 2013; Accepted October 20, 2013; Epub November 1, 2013; Published November 15, 2013
Abstract: DNA polymerase ε (polε) plays a central role in DNA replication in eukaryotic cells, and has been suggested to the
main synthetic polymerase on the leading strand. It is a hetero-tetrameric enzyme, comprising a large catalytic subunit (the A
subunit ~250 kDa), a B subunit of ~60 kDa in most species (~80 kDa in budding yeast) and two smaller subunits (each ~20
kDa). In Drosophila, two subunits of polε (dpolε) have been identified. One is the 255 kDa catalytic subunit (dpolεp255), and the
other is the 58 kDa subunit (dpolεp58). The functions of the B subunit have been mainly studied in budding yeast and
mammalian cell culture, few studies have been performed in the context of an intact multicellular organism and therefore its
functions in this context remain poorly understood. To address this we examined the in vivo role of dpolεp58 in Drosophila. A
homozygous dpolεp58 mutant is pupal lethal, and the imaginal discs are less developed in the third instar larvae. In the eye
discs of this mutant S phases, as measured by BrdU incorporation assays, were significantly reduced. In addition staining with
an anti-phospho histone H3 (PH3) antibody, (a marker of M phase), was increased in the posterior region of eye discs, where
usually cells stop replicating and start differentiation. These results indicate that dpolεp58 is essential for Drosophila
development and plays an important role in progression of S phase in mitotic cell cycles. We also observed that the size of
nuclei in salivary gland cells were decreased in dpolεp58 mutant, indicating that dpolεp58 also plays a role in endoreplication.
Furthermore we detect a putative functional interaction between dpolε and ORC2 in discs suggesting that polε plays a role in the
initiation of DNA replication in Drosophila. (ajcr0000232).
Keywords: DNA polymerase ε B subunit, Drosophila melanogaster
Address correspondence to: Masamitsu Yamaguchi, Department of Applied Biology, Kyoto Institute of Technology,
Matsugasaki, Sakyo-ku, Kyoto 606-8585, Japan. Tel: +81 75 724 7781; E-mail: myamaguc@kit.ac.jp
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American Journal of Cancer Research
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