Phosphorylation of DNA topoisomerase I is increased during the response of mammalian cells to mitogenic stimuli

D. Scott Samuels; Yoshiko Shimizu; Toshikatsu Nakabayashi; Nobuyoshi Shimizu

doi:10.1016/0167-4889(94)90075-2

Phosphorylation of DNA topoisomerase I is increased during the response of mammalian cells to mitogenic stimuli

D. Scott Samuels
, Yoshiko Shimizu
, Toshikatsu Nakabayashi
, Nobuyoshi Shimizu

Division of Biological and Biomedical Sciences

Research output: Contribution to journal › Article › peer-review

13 Scopus citations

Abstract

DNA topoisomerase I is phosphorylated after mitogenic stimulation of 3T3-L1 mouse fibroblasts by 12-O-tetradecanoylphorbol 13-acetate (TPA), a phorbol ester tumor promoter. In vivo labeling with [³²P]orthophosphate and immunoprecipitation with an anti-DNA topoisomerase I antibody has demonstrated an increase in the phosphorylation of DNA topoisomerase I in Swiss/3T3 mouse fibroblasts treated with epidermal growth factor (EGF) and H35 rat hepatoma cells treated with insulin. The only phosphorylated form of DNA topoisomerase I observed was the 100-kDa protein. Digestion of DNA topoisomerase I with trypsin revealed two phosphopeptides. In addition, VT-1, a non-responsive genetic variant of 3T3-L1, and the DNA topoisomerase I inhibitor camptothecin were used to further study TPA-induced DNA topoisomerase I phosphorylation. Our results indicate that the phosphorylation of DNA topoisomerase I may be an ubiquitous response of cultured mammalian cells to mitogenic agents, even in the absence of DNA replication.

Original language	English
Pages (from-to)	77-83
Number of pages	7
Journal	BBA - Molecular Cell Research
Volume	1223
Issue number	1
DOIs	https://doi.org/10.1016/0167-4889(94)90075-2
State	Published - Aug 11 1994

Funding

We thank Witold Cieplak and Neil Margolis for thoughtful and critical reading of the manuscript; Vas Aposhian, Witold Cieplak, Jendy Hall, Tom Lindell, John Little, and Sam Ward for valuable comments; Peter D'Arpa and Tao Hsieh for useful discussions; Mitsuo Homma and Takeshi Tojo for kindly providing purified scleroderma antibodies; Witold Cieplak and Dave Mead for HPLC analysis; and Bob Evans and Gary Hettrick for help with figure preparation. This research was supported by a Public Health Service Grant from the National Institutes of Health (GM-24375) and a Cancer Biology Predoctoral Training Award sponsored by a National Research Service Award from the National Cancer Institute (CA-09213).

Funder number
GM-24375
T32CA009213

Keywords

(Fibroblast)
(Hepatoma cell)
(Mouse)
12-O-Tetradecanoylphorbol 13-acetate
Camptothecin
DNA topoisomerase I
Epidermal growth factor
Insulin
Phosphorylation
TPA

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10.1016/0167-4889(94)90075-2

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@article{cac7a0a03c7b4f74a890c8164065ac1a,

title = "Phosphorylation of DNA topoisomerase I is increased during the response of mammalian cells to mitogenic stimuli",

abstract = "DNA topoisomerase I is phosphorylated after mitogenic stimulation of 3T3-L1 mouse fibroblasts by 12-O-tetradecanoylphorbol 13-acetate (TPA), a phorbol ester tumor promoter. In vivo labeling with [32P]orthophosphate and immunoprecipitation with an anti-DNA topoisomerase I antibody has demonstrated an increase in the phosphorylation of DNA topoisomerase I in Swiss/3T3 mouse fibroblasts treated with epidermal growth factor (EGF) and H35 rat hepatoma cells treated with insulin. The only phosphorylated form of DNA topoisomerase I observed was the 100-kDa protein. Digestion of DNA topoisomerase I with trypsin revealed two phosphopeptides. In addition, VT-1, a non-responsive genetic variant of 3T3-L1, and the DNA topoisomerase I inhibitor camptothecin were used to further study TPA-induced DNA topoisomerase I phosphorylation. Our results indicate that the phosphorylation of DNA topoisomerase I may be an ubiquitous response of cultured mammalian cells to mitogenic agents, even in the absence of DNA replication.",

keywords = "(Fibroblast), (Hepatoma cell), (Mouse), 12-O-Tetradecanoylphorbol 13-acetate, Camptothecin, DNA topoisomerase I, Epidermal growth factor, Insulin, Phosphorylation, TPA",

author = "Samuels, \{D. Scott\} and Yoshiko Shimizu and Toshikatsu Nakabayashi and Nobuyoshi Shimizu",

note = "Funding Information: We thank Witold Cieplak and Neil Margolis for thoughtful and critical reading of the manuscript; Vas Aposhian, Witold Cieplak, Jendy Hall, Tom Lindell, John Little, and Sam Ward for valuable comments; Peter D'Arpa and Tao Hsieh for useful discussions; Mitsuo Homma and Takeshi Tojo for kindly providing purified scleroderma antibodies; Witold Cieplak and Dave Mead for HPLC analysis; and Bob Evans and Gary Hettrick for help with figure preparation. This research was supported by a Public Health Service Grant from the National Institutes of Health (GM-24375) and a Cancer Biology Predoctoral Training Award sponsored by a National Research Service Award from the National Cancer Institute (CA-09213).",

year = "1994",

month = aug,

day = "11",

doi = "10.1016/0167-4889(94)90075-2",

language = "English",

volume = "1223",

pages = "77--83",

journal = "BBA - Molecular Cell Research",

issn = "0167-4889",

publisher = "Elsevier B.V.",

number = "1",

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TY - JOUR

T1 - Phosphorylation of DNA topoisomerase I is increased during the response of mammalian cells to mitogenic stimuli

AU - Samuels, D. Scott

AU - Shimizu, Yoshiko

AU - Nakabayashi, Toshikatsu

AU - Shimizu, Nobuyoshi

N1 - Funding Information: We thank Witold Cieplak and Neil Margolis for thoughtful and critical reading of the manuscript; Vas Aposhian, Witold Cieplak, Jendy Hall, Tom Lindell, John Little, and Sam Ward for valuable comments; Peter D'Arpa and Tao Hsieh for useful discussions; Mitsuo Homma and Takeshi Tojo for kindly providing purified scleroderma antibodies; Witold Cieplak and Dave Mead for HPLC analysis; and Bob Evans and Gary Hettrick for help with figure preparation. This research was supported by a Public Health Service Grant from the National Institutes of Health (GM-24375) and a Cancer Biology Predoctoral Training Award sponsored by a National Research Service Award from the National Cancer Institute (CA-09213).

PY - 1994/8/11

Y1 - 1994/8/11

N2 - DNA topoisomerase I is phosphorylated after mitogenic stimulation of 3T3-L1 mouse fibroblasts by 12-O-tetradecanoylphorbol 13-acetate (TPA), a phorbol ester tumor promoter. In vivo labeling with [32P]orthophosphate and immunoprecipitation with an anti-DNA topoisomerase I antibody has demonstrated an increase in the phosphorylation of DNA topoisomerase I in Swiss/3T3 mouse fibroblasts treated with epidermal growth factor (EGF) and H35 rat hepatoma cells treated with insulin. The only phosphorylated form of DNA topoisomerase I observed was the 100-kDa protein. Digestion of DNA topoisomerase I with trypsin revealed two phosphopeptides. In addition, VT-1, a non-responsive genetic variant of 3T3-L1, and the DNA topoisomerase I inhibitor camptothecin were used to further study TPA-induced DNA topoisomerase I phosphorylation. Our results indicate that the phosphorylation of DNA topoisomerase I may be an ubiquitous response of cultured mammalian cells to mitogenic agents, even in the absence of DNA replication.

AB - DNA topoisomerase I is phosphorylated after mitogenic stimulation of 3T3-L1 mouse fibroblasts by 12-O-tetradecanoylphorbol 13-acetate (TPA), a phorbol ester tumor promoter. In vivo labeling with [32P]orthophosphate and immunoprecipitation with an anti-DNA topoisomerase I antibody has demonstrated an increase in the phosphorylation of DNA topoisomerase I in Swiss/3T3 mouse fibroblasts treated with epidermal growth factor (EGF) and H35 rat hepatoma cells treated with insulin. The only phosphorylated form of DNA topoisomerase I observed was the 100-kDa protein. Digestion of DNA topoisomerase I with trypsin revealed two phosphopeptides. In addition, VT-1, a non-responsive genetic variant of 3T3-L1, and the DNA topoisomerase I inhibitor camptothecin were used to further study TPA-induced DNA topoisomerase I phosphorylation. Our results indicate that the phosphorylation of DNA topoisomerase I may be an ubiquitous response of cultured mammalian cells to mitogenic agents, even in the absence of DNA replication.

KW - (Fibroblast)

KW - (Hepatoma cell)

KW - (Mouse)

KW - 12-O-Tetradecanoylphorbol 13-acetate

KW - Camptothecin

KW - DNA topoisomerase I

KW - Epidermal growth factor

KW - Insulin

KW - Phosphorylation

KW - TPA

UR - https://www.scopus.com/pages/publications/0027964647

U2 - 10.1016/0167-4889(94)90075-2

DO - 10.1016/0167-4889(94)90075-2

M3 - Article

C2 - 8061056

AN - SCOPUS:0027964647

SN - 0167-4889

VL - 1223

SP - 77

EP - 83

JO - BBA - Molecular Cell Research

JF - BBA - Molecular Cell Research

IS - 1

ER -

Phosphorylation of DNA topoisomerase I is increased during the response of mammalian cells to mitogenic stimuli

Abstract

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Keywords

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