초록

Ⅳ. 연구개발결과
- 개구리 난자를 통한 체세포 리프로그래밍 시스템을 구축
- 핵치환 리프로그래밍 기술에 사용되는 MII 난자의 새로운 세포주기 조절 기전 규명
- 개구리 난자 추출액 분활화를 통한 리프로그래밍 인자 발굴
- 데이터베이스 분석을 통한 리프로그래밍 인자 발굴

Abstract ▼

DNAs from sperm are highly condensed and transcriptionally inert. After fertilization, sperm DNAs start to be decondensed and reorganized to initiate transcription, which is essential for normal development. Considering that sperm only provide DNA to start life, it is reasonable that eggs and oocyte

DNAs from sperm are highly condensed and transcriptionally inert. After fertilization, sperm DNAs start to be decondensed and reorganized to initiate transcription, which is essential for normal development. Considering that sperm only provide DNA to start life, it is reasonable that eggs and oocytes possess all components essential for development. These components include intrinsic factors to induce transcription of sperm after fertilization. Recent studies showed that such a remarkable event that occurs after fertilization takes also place at the somatic cells after somatic nuclei are transplanted into enucleated eggs or oocytes. Therefore,eggs and oocytes seem to have reprogramming factors to induce the transcription of silenced genes including Oct4 and Sox2.
Reprogramming factors are synthesized and accumulated during oocyte maturation and are especially in the nucleus, called the germinal vesicle (GV). It is unclear what kinds of GV factors are required for reprogramming and for normal development, and how they contribute to these fundamental processes. In this study, we have developed a nuclear transfer assay to identify such a maternal factor. Firstly, we injected hundreds of mammalian somatic cell nuclei into the GV of Xenopus oocytes and the transcription of Oct4/Sox2 was determined. We found that pluoripotency genes including Oct4/Sox2 are reactivated within 48 hours after transfer.Therefore, this nuclear transfer system provides a unique opportunity to identify maternal factors responsible for reprogramming the transcription of somatic nuclei. Secondly, we have treated the permeabilized somatic cells with the egg extracts and found that the reprogramming has been occurred in vitro condition,
suggesting that egg extracts contain sufficient amounts of maternal factors and these factors are biologically active in vitro condition. To identify the maternal factors, therefore, we have fractionated the extracts and tested the ability to express the Oct4/Sox2 after subtraction of a single fraction. We are currently conducting this experiment to identify the maternal factors. As an alternative approach, we have screened Oct4-binding partners by computer analyses and constructed Oct4-interactome database. These database will broaden our understanding on the stem cell biology. Lastly, we also found that the fundamental mechanism that regulates cell cycle resumption after fertilization, a prerequisite for reprogramming, is regulated either by Cdc25A or Greatwall kinase.