Ouellette A J, Reed S L, Malt R A
Proc Natl Acad Sci U S A. 1976 Aug;73(8):2609-13. doi: 10.1073/pnas.73.8.2609.
In experiments originally designed to examine selective turnover of methylated "caps" in renal mRNA, we observed that [3H]methyl label decayed from mRNA containing poly(A) with a half-life of 1-2 hr. (Caps are blocked, methylated mRNA sequences of the general structure m7GpppNm p(1 or 2)Np.). To distinguish between metabolism of short-lived mRNA and discriminate turnover of "caps", we compared residual [3H]methyl label in 5' and 3'mRNA fragments prepared from mRNA isolated during the decay period. Hydrolysis of mRNA at 0 degrees with dilute KOH before oligo(dT)-cellulose selection produced 5' mRNA fragments enriched with an alkali-resistant oligonucleotide with a -5 charge; the 3' mRNA fraction was correspondingly reduced in oligonucleotide content. Since methyl label disappeared at the same rate from both fractions, we conclude that mouse kidney contains short-lived mRNA and that the "caps" of these labile mRNAs turn over with the rest of the mRNA molecule.
在最初设计用于检测肾mRNA中甲基化“帽”的选择性周转的实验中,我们观察到[3H]甲基标记从含有聚(A)的mRNA中衰减,半衰期为1 - 2小时。(帽是一般结构为m7GpppNm p(1或2)Np的封闭、甲基化的mRNA序列。)为了区分短寿命mRNA的代谢和“帽”的差异周转,我们比较了在衰变期从分离的mRNA制备的5'和3'mRNA片段中的残留[3H]甲基标记。在寡聚(dT)-纤维素选择之前,用稀KOH在0摄氏度水解mRNA产生富含带-5电荷的耐碱寡核苷酸的5'mRNA片段;3'mRNA部分的寡核苷酸含量相应减少。由于甲基标记从两个部分以相同的速率消失,我们得出结论,小鼠肾脏含有短寿命mRNA,并且这些不稳定mRNA的“帽”与mRNA分子的其余部分一起周转。