Identification of radiation-sensitive mutants in the Medaka, Oryzias latipes.
We screened populations of N-ethyl-N-nitrosourea (ENU)-mutagenized Medaka, (Oryzias latipes) for radiation-sensitive mutants to investigate the mechanism of genome stability induced by ionizing radiation in developing embryos. F3 embryos derived from male founders that were homozygous for induced the mutations were irradiated with gamma-rays at the organogenesis stage (48hpf) at a dose that did not cause malformation in wild-type embryos. We screened 2130 F2 pairs and identified three types of mutants with high incidence of radiation-induced curly tailed (ric) malformations using a low dose of irradiation. The homozygous strain from one of these mutants, ric1, which is highly fertile and easy to breed, was established and characterized related to gamma-irradiation response. The ric1 strain also showed higher incidence of malformation and lower hatchability compared to the wild-type CAB strain after gamma-irradiation at the morula and pre-early gastrula stages. We found that the decrease in hatching success after gamma-irradiation, depends on the maternal genotype at the ric1 locus. Terminal deoxynucleotidyl transferase-mediated deoxy-UTP nick end-labeling assays showed a high frequency of apoptosis in the ric1 embryos immediately after gamma-irradiation at the pre-early gastrula stage but apoptotic cells were not observed before midblastula transition (MBT). The neutral comet assay revealed that the ric1 mutant has a defect in the rapid repair of DNA double-strand breaks induced by gamma-rays. These results suggest that RIC1 is involved in the DNA double strand break repair in embryos from morula to organogenesis stages, and unrepaired DNA double strand breaks in ric1 trigger apoptosis after MBT. These results support the use of the ric1 strain for investigating various biological consequences of DNA double strand breaks in vivo and for sensitive monitoring of genotoxicity related to low dose radiation.
Aizawa
Kouichi
K
Department of Integrated Biosciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa-no-ha 5-1-5, Kashiwa, Chiba 277-8572, Japan.
Mitani
Hiroshi
H
Kogure
Nozomi
N
Shimada
Atsuko
A
Hirose
Yukihiro
Y
Sasado
Takao
T
Morinaga
Chikako
C
Yasuoka
Akihito
A
Yoda
Hiroki
H
Watanabe
Tomomi
T
Iwanami
Norimasa
N
Kunimatsu
Sanae
S
Osakada
Masakazu
M
Suwa
Hiroshi
H
Niwa
Katsutoshi
K
Deguchi
Tomonori
T
Hennrich
Thorsten
T
Todo
Takeshi
T
Shima
Akihiro
A
Kondoh
Hisato
H
Furutani-Seiki
Makoto
M
D000818:Animals; D017209:Apoptosis / Q000528:radiation effects; D020552:Comet Assay; D004260:DNA Repair / Q000502:physiology; D005720:Gamma Rays; D005775:Gastrula / Q000502:physiology; D042822:Genomic Instability; D009154:Mutation; D009990:Oryzias / Q000196:embryology / Q000235:genetics*; D011836:Radiation Tolerance / Q000235:genetics*; D013623:Tail / Q000002:abnormalities / Q000196:embryology / Q000528:radiation effects; D013997:Time Factors