Changes in mutagenesis sensitivity with maturation of male germ cells as revealed by the Medaka specific-locus test system

本研究的目的是详细阐明男性生殖细胞在精子发生和精子发生过程中诱变敏感性的变化。为此，使用了Shima和Shimada开发的Medaka特定位点测试系统。所使用的遗传终点是显性致死突变、全特定位点突变和存活特定位点突变。野生型雄性青竹被伽马射线诱变。允许它们与未处理的雌性受试者交配，这些雌性受试者在三个标记座位上为纯合子隐性。在显性致死效应方面，暴露后1~3d，精子阶段的雄性生殖细胞诱发率最高，随后在暴露后第30天，诱发率单调下降，表明突变与成熟有很高的相关性。这一趋势也适用于总的特定基因座突变以及可存活的特定基因座突变。因此，伽玛射线诱发的雄性生殖细胞显性致死突变和特定位点突变的变化与从干细胞到成熟精子的雄性生殖细胞的成熟(分化)几乎是平行的。为了了解这些遗传终点背后的分子机制，提取了个体显性致死胚胎的基因组DNA，并用AP-PCR技术制备了DNA指纹。结果发现，与正常同胞相比，指纹的DNA条带丢失的发生率更高。

The purpose of the present study is to elucidate in detail the changes in mutagenesis sensitivity of male germ cells during spermatogenesis and spermiogenesis as well. For this purpose, the Medaka specific-locus test system developed by Shima and Shimada was used. The genetic endpoints employed were dominant lethal mutations, total specific-locus mutations and viable specific-locus mutations. The wild type male Medaka were mutagenized with gamma-rays. They were allowed to mate with non-treated tester females which were homozygous recessive at the three marker loci. Regarding the induction of dominant lethals with gamma-rays, the rates were highest during 1-3 days after exposure corresponding to the exposure of male germ cells at the stage of spermatozoa, followed by monotonous decrease toward 30th day after male exposure, indicating a high correlation between mutability and maturation. This trend was also the case for the total specific-locus mutations as well as viable specific-locus mutations. It was concluded, therefore, that the changes in mutability induced with gamma-rays in terms of dominant lethals and specific-locus mutations in the male germ cells run almost in parallel with the maturation (differentiation) of the male germ cells ranging from stem spermatogonia to mature spermatozoa. To understand the molecular mechanisms underlying these genetic endpoints, the genomic DNA of individual dominant lethal embryos were extracted and the DNA fingerprints were prepared by using AP-PCR.It was found that the rate of DNA alterations in terms of band loss from the fingerprint is more freuent compared to that in the normal sibling.