INOSITOL POLYPHOSPHATES ARE NOT INCREASED BY OVEREXPRESSION OF INS(1,4,5)P-3 3-KINASE BUT SHOW CELL-CYCLE DEPENDENT CHANGES IN GROWTH FACTOR-STIMULATED FIBROBLASTS
INOSITOL POLYPHOSPHATES ARE NOT INCREASED BY OVEREXPRESSION OF INS(1,4,5)P-3 3-KINASE BUT SHOW CELL-CYCLE DEPENDENT CHANGES IN GROWTH FACTOR-STIMULATED FIBROBLASTS
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DOI：
10.1091/mbc.5.1.17
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发表时间：
1994-01-01
影响因子：
3.3
通讯作者：
CATT, KJ
中科院分区：
文献类型：
--
作者：
BALLA, T;SIM, SS;CATT, KJ
将大鼠脑肌醇1，4，5-三磷酸（Ins（1，4，5）P-3）3-激酶稳定转染NIH 3 T3成纤维细胞，探讨Ins（1，3，4，5）P-4的产生与InsP（5）和InsP（6）形成的关系。InsP（5）和InsP（6）的质量测量显示激酶转染和载体转染的成纤维细胞之间没有显著差异。然而，当用[H-3]肌醇标记48-72小时时，这种3-激酶转染的细胞显示出比它们的载体转染的对应物更低的[H-3]InsP（5）和[H-3]InsP（6）以及[H-3] Ins（1，3，4，6）P-4和D/L[H-3]Ins（1，4，5，6）P-4水平。由于转染Ins（1，4，5）P-33-激酶的细胞生长速度低于载体转染的对照组，我们确定InsP（5）和InsP（6）的合成是否与细胞周期的特定阶段有关。当用[H-3]肌醇预标记的NIH 3 T3细胞通过血清剥夺和血小板衍生生长因子（PDGF）刺激进行同步化时，标记的InsP（5）和InsP（6）的量仅在刺激12 h后开始增加，此时细胞进入S期，如[H-3]胸苷掺入增加所示。这些肌醇多磷酸的合成增强之前，Ins（1，4，5）P-3及其代谢产物的早期增加，这在PDGF作用的第5小时不再明显。D/L-Ins（1，4，5，6）P-4水平也有明显的双相性升高，在3 h左右出现一个早期峰值，随后出现第二次升高，与InsP（5）和InsP（6）的升高相一致。这些结果表明，高度磷酸化肌醇的形成并不紧密耦合到受体介导的形成的Ins（1，4，5）P-3及其代谢产物，但主要是由其他因素，在细胞周期的特定点操作。
NIH 3T3 fibroblasts were stably transfected with rat brain inositol 1,4,5-trisphosphate (Ins(1,4,5)P-3) 3-kinase to explore the relationship between increased production of Ins(1,3,4,5)P-4 and the formation of InsP(5) and InsP(6). Mass measurements of InsP(5) and InsP(6) revealed no significant difference between kinase- and vector-transfected fibroblasts. However, such 3-kinase-transfected cells, when labeled with [H-3]inositol for 48-72 h, showed lower levels of [H-3]InsP(5) and [H-3]InsP(6), as well as [H-3]Ins(1,3,4,6)P-4 and D/L[H-3]Ins(1,4,5,6)P-4, than their vector-transfected counterparts. Because Ins(1,4,5)P-3 3-kinase-transfected cells grew less rapidly than vector-transfected controls, we determined whether the synthesis of InsP(5) and InsP(6) was related to a specific phase of the cell cycle. When NIH 3T3 cells prelabeled with [H-3]inositol were synchronized by serum deprivation followed by stimulation with platelet-derived growth factor (PDGF), the amounts of labeled InsP(5) and InsP(6) began to increase only after 12 h of stimulation, when cells entered the S-phase as indicated by increased [H-3]thymidine incorporation. The enhanced synthesis of these inositol polyphosphates was preceded by an early increase in Ins(1,4,5)P-3 and its metabolites that was no longer evident by the fifth hour of PDGF action. There was also a prominent and biphasic increase in the level of D/L-Ins(1,4,5,6)P-4 with an early peak at similar to 3 h and a second rise that paralleled the increases in InsP(5) and InsP(6). These results indicate that the formation of highly phosphorylated inositols is not tightly coupled to the receptor-mediated formation of Ins(1,4,5)P-3 and its metabolites but is mainly determined by other factors that operate at specific points of the cell cycle.