BIOCHEMICAL-STUDIES OF MAMMALIAN OOGENESIS - SYNTHESIS AND STABILITY OF VARIOUS CLASSES OF RNA DURING GROWTH OF THE MOUSE OOCYTE INVITRO
BIOCHEMICAL-STUDIES OF MAMMALIAN OOGENESIS - SYNTHESIS AND STABILITY OF VARIOUS CLASSES OF RNA DURING GROWTH OF THE MOUSE OOCYTE INVITRO
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DOI：
10.1016/0012-1606(81)90195-0
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发表时间：
1981-01-01
影响因子：
2.7
通讯作者：
SCHULTZ, RM
中科院分区：
文献类型：
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作者：
BROWER, PT;GIZANG, E;SCHULTZ, RM
在含有放射性标记尿苷的培养基中孵育卵泡后，检测了不同生长阶段小鼠卵母细胞中各种RNA的合成。放射性标记的卵母细胞RNA在聚（U）-琼脂糖上分级分离后，其中约83%在5小时标记期后与细胞核结合，显示约40-50%的放射性标记的RNA表现为含聚（A）的RNA。在卵母细胞生长期间，该值保持相当恒定，其中卵母细胞直径从0.4增加。35至. apprx。55 μ m标记5 h后，在完全发育的网状卵母细胞、中期II卵母细胞或1-细胞胚胎中，放射性标记的含poly（A）的RNA的百分比约为20%。标记5 h后，对在聚（U）-琼脂糖凝胶上分级分离后获得的放射性标记的卵母细胞RNA进行琼脂糖凝胶电泳分析，发现存在推定的核糖体RNA前体、核糖体（28和18 S）RNA、转移加5 S RNA和异源分散的含聚（A）RNA。放射性标记的RNA种类的显著部分相当大（> 40 S）。放射性标记的rRNA和转移加5S RNA的相对比例的比率在卵母细胞生长过程中基本保持恒定。通过用放射性标记的尿苷孵育卵泡，洗涤卵泡使其不含放射性，并在支持卵母细胞体外生长的条件下培养卵泡，检查各类RNA的稳定性。在这些条件下，通过酸不溶性放射性物质的保留测定，总卵母细胞放射性标记RNA相当稳定（t1/2 [半衰期] = 28天）。在卵母细胞有活力但不生长的条件下，总RNA的半衰期约为4.5天。含多聚腺苷酸的RNA也非常稳定;培养8天后，标记5小时后仍存在约50%的放射性标记的含多聚腺苷酸的RNA。培养4天后和在聚（U）-琼脂糖凝胶上分级分离后，对卵母细胞中放射性标记的RNA进行琼脂糖凝胶电泳分析，发现存在核糖体（28和18 S）RNA、转移加5 S RNA和异源分散的含聚（A）RNA。此时，这些RNA位于卵母细胞的细胞质中。此外，含有poly（A）的RNA的MW分布显著低于标记5 h后的MW分布。放射性标记的r RNA和转移加5 S RNA的相对比例的比值与标记5 h后的比值非常相似。
The synthesis of various classes of RNA in mouse oocytes at different stages of growth was examined after incubating follicles in medium containing radiolabeled uridine. After fractionation on poly(U)-Sepharose of radiolabeled oocyte RNA, of which about 83% is associated with the nucleus after a 5 h labeling period, revealed that about 40-50% of the radiolabeled RNA behaved as poly(A)-containing RNA. This value remained fairly constant during the period of oocyte growth in which oocyte diameter increased from .apprx. 35 to .apprx. 55 .mu.m. After a 5 h labeling, the percentage of radiolabeled poly(A)-containing RNA in either the fully grown dictyate oocyte, metaphase II oocyte or 1-cell embryo was about 20%. After a 5 h labeling, agarose gel electrophoretic analysis of the radiolabeled species of oocyte RNA obtained after fractionation on poly(U)-Sepharose revealed the presence of putative ribosomal RNA precursor, ribosomal (28 and 18 S) RNA, transfer plus 5 S RNA and heterodisperse poly(A)-containing RNA. A significant fraction of the radiolabeled RNA species was quite large (> 40 S). The ratios of the relative proportions of the radiolabeled rRNA and transfer plus 5 S RNA remained essentially constant during oocyte growth. The stability of various classes of RNA was examined by incubating follicles with radiolabeled uridine, washing the follicles free of radioactivity and culturing the follicles under conditions which support oocyte growth in vitro. Under these conditions, total oocyte radiolabeled RNA was quite stable as determined by retention of acid-insoluble radioactive material (t1/2 [half-life] = 28 days). Under conditions in which oocytes are viable but do not grow, the half-life of total RNA was about 4.5 days. Poly(A)-containing RNA was also very stable; after 8 days in culture, about 50% of the radiolabeled poly(A)-containing RNA present after 5 h of labeling was still present. Agarose gel electrophoretic analysis of radiolabeled RNA in oocytes after 4 days of culture and after fractionation on poly(U)-Sepharose revealed the presence of ribosomal (28 and 18 S) RNA, transfer plus 5 S RNA, and heterodisperse poly(A)-containing RNA. At this time, these RNA are located in the oocyte cytoplasm. In addition, the MW distribution of poly(A)-containing RNA was significantly lower than that after 5 h of labeling. The ratios of the relative proportions of radiolabeled r RNA and transfer plus 5 S RNA were quite similar to those after 5 h of labeling.