논문 상세보기
pp.75-85
생쥐 초기 배아의 형태형성에 영향을 주는 세포질내 인자의 기원과 작용기작을 연구하기 위해 단백질 합성과 단백질 활성화 효소 (protein kinase)의 억제제를 처리한 배아의 세포질로 재조합된 배아에서 발생과 RNA합성, 단백질 인산화를 조사하였다. 단백질 합성 억제제인 cycloheximide (CHX)가 함유된 배양액에서 24시간 배양한 1-세포 배아의 제핵된 세포질을 두 개의 전핵을 모두 가진 절반의 1-세포 배아와 재조합한 P+P-CHX군의
To investigate the origin and action mechanism of cytoplasmic factors as regulators of morphogenesis, the embryonic development, RNA synthesis and protein phosphorylation were examined in reconstituted embryos. A half of 1-cell mouse embryo with both pronuclei was electrofused with the enucleated cytoplasm of 1- or 2-cell embryos which were cultured for 24 hrs from post 20 hrs hCG in CZB with or without cycloheximide (CHX, an inhibitor of protein synthesis; P+P-CHX group), genistein (Gen, an inhibitor of tyrosine protein kinase; P+2-Gen group) and 6-dimethylaminopurine (6-DMAP, an inhibitor of serine-threonine protein kinase; P＋2-DMAP group), and co-cultured with Vero cells for 5 days. And their development, cell numbers at compaction, [5, 6-H]-uridine incorporation into RNA and the pattern of protein phosphorylation after labeling of [ P] orthophosphate were compared with that of the reconstituted embryos such as P+2 or P+P (control group). Embryonic development and the time of RNA synthesis in P+P-CHX were similar to those in P+P. But the time and the cell stages of embryonic compaction in P+P-CHX were similar to those in P+2. The compaction was initiated at 4-cell in P+2 and P+2-Gen, but at 8-cell in P+P and P+2-DMAP. On a two-dimensional gel electrophoresis, phosphorylation of 80KD and 110KD proteins were inhibited after 3 hrs of reconstruction in the embryo of P+2-DMAP when compared with that of P+2 and P+2-Gen. These results suggest that protein synthesis between 1- and 2-cell stage affects the timing of embryonic genome activation, and that cytoplasmic factors derived from oocyte or their modification regulates the time schedule of embryonic compaction in mouse. Also, serine-threonine protein kinase has an important role on the regulation of compaction.
