Williamson J D, Quatrano R S, Cuming A C
Eur J Biochem. 1985 Oct 15;152(2):501-7. doi: 10.1111/j.1432-1033.1985.tb09224.x.
The effect of abscisic acid (ABA) on the expression of the 'early-methionine-labeled' (Em) polypeptide was examined in cultured, immature wheat (Triticum aestivum, L.) embryos and in developing embryos in planta. A complementary DNA (cDNA) library was constructed from poly(A)-rich RNA from immature embryos cultured in the presence of ABA. ABA-enhanced sequences were first identified by differential colony-blot hybridization, and then verified using RNA slot-blot analysis. Dot-blot hybridization showed that one clone, p1015, was homologous to the previously isolated Em cDNA, pWG432. Electrophoretic analysis of the hybrid-select translation product of p1015 confirmed its identity as an Em sequence. Comparison of the p1015 cDNA insert size and the Em message size, from northern blot analysis, showed that p1015 contained about 87% of the Em sequence. RNA slot-blot analysis and protein electrophoresis showed that Em message, but not Em protein, accumulated at a low, basal level in immature embryos in the absence of ABA. Neither Em message nor Em protein was seen in three-day germinated seedlings. Steady-state levels of Em message and protein increased in immature embryos in the presence of ABA, both in culture and in planta. Regulation appeared to be primarily at the level of transcription or specific message stability. Regulation may also involve specific protein stability, since synthesis of Em protein continued in immature embryos in the absence of ABA, but Em protein did not accumulate in detectable amounts. We conclude that ABA specifically modulates Em message and protein levels in immature embryos, but is probably not responsible for the embryogenic specificity of Em expression.
在培养的未成熟小麦（Triticum aestivum, L.）胚以及植株中发育的胚中，研究了脱落酸（ABA）对“早期甲硫氨酸标记”（Em）多肽表达的影响。从在ABA存在下培养的未成熟胚富含多聚腺苷酸（poly(A)）的RNA构建了一个互补DNA（cDNA）文库。首先通过差异菌落杂交鉴定出ABA增强的序列，然后使用RNA狭缝印迹分析进行验证。斑点印迹杂交表明，一个克隆p1015与先前分离的Em cDNA pWG432同源。对p1015的杂交选择翻译产物进行电泳分析，证实其为Em序列。通过Northern印迹分析比较p1015 cDNA插入片段大小和Em信息大小，表明p1015包含约87%的Em序列。RNA狭缝印迹分析和蛋白质电泳表明，在没有ABA的情况下，未成熟胚中Em信息以低基础水平积累，但Em蛋白不积累。在三天发芽的幼苗中既未检测到Em信息也未检测到Em蛋白。在ABA存在下，无论是在培养物中还是在植株中，未成熟胚中Em信息和蛋白质的稳态水平均升高。调控似乎主要在转录或特定信息稳定性水平。调控也可能涉及特定蛋白质的稳定性，因为在没有ABA的情况下，未成熟胚中Em蛋白的合成仍在继续，但Em蛋白没有积累到可检测的量。我们得出结论，ABA特异性调节未成熟胚中Em信息和蛋白质水平，但可能不负责Em表达的胚胎发生特异性。
