Watson A J, Pape C, Emanuel J R, Levenson R, Kidder G M
Department of Zoology, University of Western Ontario, London, Canada.
Dev Genet. 1990;11(1):41-8. doi: 10.1002/dvg.1020110106.
Na,K-ATPase is a plasma membrane enzyme that plays a critical role in eutherian blastocoel formation (cavitation) by pumping Na+ into the extracellular space enclosed by the trophectoderm. Previous experiments with the mouse had shown that the alpha (catalytic) subunit of the enzyme becomes detectable by immunocytochemistry in the late morula, just prior to the onset of cavitation. In the present study we have used cDNAs corresponding to three mRNA isoforms of the alpha subunit and a beta subunit to determine which genes are expressed during preimplantation development and to explore the timing of their expression. Of the three alpha subunit cDNAs tested by Northern blot hybridization with blastocyst RNA, only alpha 1 produced a hybridization signal, recognizing a single mRNA about 4 kb in length. This mRNA is relatively abundant in zygotes but barely detectable by the 2-cell stage and then accumulates steadily thereafter to reach its preimplantation maximum in blastocysts. The beta 1 cDNA detected mRNA of about 2.6-2.8 kb. This mRNA is present in zygotes but could not be detected in 2-, 4-, or 8-cell stages; it is present at a low level in late morulae and is abundant in blastocysts. The temporal profile of accumulation of beta 1 mRNA thus matches more closely than does alpha 1 the timing of appearance of the catalytic subunit. This suggests that the beta subunit may regulate production of the holoenzyme and hence the timing of cavitation.
钠钾ATP酶是一种质膜酶,通过将钠离子泵入滋养外胚层所包围的细胞外空间,在真兽类囊胚腔形成(空化)过程中发挥关键作用。先前对小鼠的实验表明,该酶的α(催化)亚基在桑椹胚晚期、即将开始空化之前,可通过免疫细胞化学检测到。在本研究中,我们使用了与α亚基的三种mRNA亚型和一个β亚基相对应的cDNA,以确定哪些基因在植入前发育过程中表达,并探索它们的表达时间。通过与囊胚RNA进行Northern印迹杂交测试的三种α亚基cDNA中,只有α1产生了杂交信号,识别出一种约4 kb长的单一mRNA。这种mRNA在合子中相对丰富,但在2细胞期几乎检测不到,此后稳步积累,在囊胚中达到植入前的最大值。β1 cDNA检测到约2.6 - 2.8 kb的mRNA。这种mRNA存在于合子中,但在2细胞、4细胞或8细胞期无法检测到;它在桑椹胚晚期含量较低,在囊胚中丰富。因此,β1 mRNA积累的时间模式比α1更紧密地匹配催化亚基出现的时间。这表明β亚基可能调节全酶的产生,从而调节空化的时间。