4-THIOURIDINE PHOTOSENSITIZED RNA-PROTEIN CROSS-LINKING IN MAMMALIAN-CELLS
4-THIOURIDINE PHOTOSENSITIZED RNA-PROTEIN CROSS-LINKING IN MAMMALIAN-CELLS
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DOI：
10.1016/s0006-291x(86)80250-9
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发表时间：
1986-12-15
影响因子：
3.1
通讯作者：
SALET, C
中科院分区：
文献类型：
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作者：
FAVRE, A;MORENO, G;SALET, C
猴肾细胞（CV-1）在存在0.1 mM 4-硫氨胺（S4U）的情况下培养，随后在365 nm处释放，表现出明显的RNA合成抑制作用。获得4 H S4U孵育和45 kJ/m2剂量的最大效应（.Apprxeq。40％）。在这些条件下，在苯酚 - 氯仿提取过程中，最多可在相间中保留了总细胞RNA的20％。与蛋白质交联的RNA分数相当于S4U掺入过程中合成的RNA标记的3H-尿素的50％，对于对照样品而言少于10％。这强烈表明，S4U掺入RNA链中的S4U充当光性探针。上面的数据提供了研究在非破坏性条件下体内RNA - 蛋白质相互作用的方法的基础。
Monkey kidney cells (CV-1) cultivated in the presence of 0.1 mM 4-thiouridine (S4U) and subsequently illuminated at 365 nm exhibit a marked RNA synthesis inhibition. Maximal effect (.apprxeq. 40%) was obtained for a 4 h S4U incubation and a 45 kJ/m2 dose. Under these conditions up to 20% of total cellular RNA is retained at the interphase during phenol-chloroform extraction. The fraction of RNA crosslinked to proteins amounts to 50% of the 3H-uridine labeled RNA synthesized during S4U incorporation and less than 10% for the control samples. This strongly suggests that S4U incorporated within the RNA chains acts as a photoaffinity probe. The data above provide the basis of a method of studying in vivo RNA-protein interactions under non destructive conditions.