INDUCTION OF AN EXTRACELLULAR RIBONUCLEASE IN CULTURED TOMATO CELLS UPON PHOSPHATE STARVATION
INDUCTION OF AN EXTRACELLULAR RIBONUCLEASE IN CULTURED TOMATO CELLS UPON PHOSPHATE STARVATION
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DOI：
10.1104/pp.92.4.970
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发表时间：
1990-04-01
期刊：
影响因子：
7.4
通讯作者：
GLUND, K
中科院分区：
文献类型：
--
作者：
NURNBERGER, T;ABEL, S;GLUND, K
番茄（Lycopersicon esculentum）悬浮培养细胞在从对数生长期向稳定生长期过渡期间开始分泌RNA降解酶活性。使用琼脂糖-5-（4-氨基苯基-磷酰基）尿苷3′（2′）单磷酸作为强有力的最终富集步骤，将酶纯化至均一，并根据以下数据表征为核糖核酸酶I（RNase I）：（a）它有一个22，000先生（十二烷基硫酸钠-聚丙烯酰胺凝胶电泳），最适pH为pH5.5，pI为3.9，对EDTA不敏感;（B）该酶通过磷酸转移酶反应以核酸内切方式裂解单链RNA，产生2′，3 ′-cNMP作为主要单体产物;（c）如用二核糖核苷单磷酸作为底物所研究的，该酶表现出对邻近裂解位点的5′嘌呤残基的明显偏好。最有趣的是，在体内合成和分泌被发现时，番茄细胞被专门饥饿的磷酸盐作为矿物质营养诱导。(a)细胞外酶活性增加约10倍后，转移到培养基中缺乏磷酸盐的磷酸盐生长的细胞。因此，当细胞持续供应磷酸盐时，这种活性的增加是检测不到的。(b)用放射性氨基酸生物合成标记的细胞外蛋白质可通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳/荧光法直接在大量细胞外蛋白质内检测到。因此，我们建议，分泌番茄核糖核酸酶I合成磷酸饥饿是一个更高的植物诱导的救援系统清除外源磷的组成部分。
Suspension-cultured cells of tomato (Lycopersicon esculentum) start to secrete an RNA-degrading enzyme activity during transition from logarithmic to stationary growth phase. Using affinity chromatography on agarose-5-(4-aminophenyl-phosphoryl) uridine 3′(2′) monophosphate as a powerful and final enrichment step, the enzyme was purified to homogeneity and characterized as ribonuclease I (RNase I) according to the following data: (a) it has anMrof 22,000 (sodium dodecyl sulfate-polyacrylamide gel electrophoresis), a pH-optimum of pH 5.5, a pl of 3.9, and its activity was found to be insensitive to EDTA; (b) the enzyme splits single-stranded RNA endonucleolytically by a phosphotransferase reaction yielding 2′,3′-cNMPs as primary monomeric products; (c) as studied with diribonucleoside monophosphates as substrates, the enzyme exhibits a pronounced preference for 5′ purine residues adjacent to the cleavage site. Most interestingly,in vivosynthesis and secretion was found to be induced when tomato cells were specifically starved for phosphate as mineral nutrient. (a) Extracellular enzyme activity increased about tenfold after transfer of phosphate-grown cells into medium lacking only phosphate. Accordingly, this increase in activity was not detectable when cells were constantly supplied with phosphate. (b) Biosynthetically labeling of the extracellular protein with radioactive amino acids was detectable by sodium dodecyl sulfate-polyacrylamide gel electrophoresis/fluorography directly within the bulk of extracellular proteins. Therefore, we propose that the secreted tomato RNase I synthesized upon phosphate starvation is a component of a higher plant inducible rescue system for scavenging exogenous phosphate.