Messenger RNA stability in Dictyostelium discoideum: does poly(A) have a regulatory role?
Messenger RNA stability in Dictyostelium discoideum: does poly(A) have a regulatory role?
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盘基网柄菌中信使 RNA 的稳定性：poly(A) 是否具有调节作用？
DOI：
10.1016/0022-2836(80)90379-4
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发表时间：
1980
影响因子：
5.6
通讯作者：
Jacobson,A
中科院分区：
文献类型：
--
作者：
Palatnik,CM;Storti,RV;Capone,AK;Jacobson,A
在先前的实验中（Palatniket等人，1979）我们用poly（U）-Sepharose热洗脱法将盘状网柄藻的信使RNA分离成poly（A）链长度不同的几个组分，这些组分的体外翻译表明，有些细胞mRNA在poly（A）链较长的组分中相对更丰富（110至115个核苷酸）和其他细胞mRNA在具有短聚（A）片段（60至65个核苷酸）的那些级分中更丰富。我们以前的实验也表明，植物生长细胞的大多数mRNA是用长的poly（A）束合成的，随着细胞的老化，这些长的poly（A）束缩短到60到65个核苷酸的稳定状态大小。本文的实验研究了这样一种可能性，即mRNA分别富集在长和短poly（A）组分中，代表不同的mRNA稳定性类别，也反映了poly（A）参与决定mRNA稳定性。用完全抑制RNA合成的浓度的放线菌素D培养盘基网柄藻的营养生长细胞。18小时后，提取细胞总RNA，在依赖信使RNA的网织红细胞裂解液中进行翻译，用二维聚丙烯酰胺凝胶电泳分析体外翻译产物，并将电泳图谱与以前分析不同poly（A）含量的mRNA所得图谱进行比较。我们的结果表明，放线菌素处理后：（a）mRNA群体的翻译复杂性显著降低;（B）至少有66种主要mRNA的相对丰度显著增加或减少;（c）相对丰度增加的大多数mRNA是那些在未处理细胞中最丰富的短poly（A）组分;和（d）相对丰度降低的大多数mRNA是那些在未处理细胞中在长poly（A）级分中最丰富的mRNA。这些实验，再加上我们以前的结果，使我们建议，那些mRNA富含含有短，稳态大小的poly（A）束的馏分是最稳定的。我们的数据不支持poly（A）长度调节mRNA稳定性的模型。1979）mRNA类，其仅包含寡核苷酸（A）片段（约。25个核苷酸）在放线菌素D的存在下也相对不稳定;和（B）在细胞脱离长期暴露于放线菌素D后，可翻译mRNA的分布发生重大变化。许多已被放线菌素D处理大幅减少的mRNA在去除药物后增加了相对丰度。此外，在放线菌素D处理的细胞中积累到非常高水平的大部分mRNA在药物去除后相对丰度急剧下降。
In previous experiments (Palatniket al., 1979) we used thermal elution from poly(U)-Sepharose to separate messenger RNA fromDictyostelium discoideuminto several fractions differing in the length of their poly(A) tracts.In vitrotranslation of these fractionated RNAs demonstrated that some cellular mRNAs are relatively more abundant in those fractions with long poly(A) tracts (110 to 115 nucleotides) and other cellular mRNAs are more abundant in those fractions with short poly(A) tracts (60 to 65 nucleotides). Our previous experiments also showed that most mRNAs of vegetatively growing cells are synthesized with long poly(A) tracts which shorten, with age in the cell, to a steady-state size of 60 to 65 nucleotides. The experiments of this paper examined the possibility that the mRNAs enriched, respectively, in the long and short poly(A) fractions, represented different stability classes of mRNA and also reflected the involvement of poly(A) in determining mRNA stability. Vegetatively growing cells ofDictyostelium discoideumwere incubated with actinomycin D at a concentration which totally inhibited synthesis of RNA. After 18 hours, total cellular RNA was extracted and translated in a messenger RNA-dependent reticulocyte lysate.In vitrotranslation products were analyzed by two-dimensional polyacrylamide gel electrophoresis and electrophoretic patterns were compared with those obtained from similar, previous analyses of mRNAs of different poly(A) content. Our results showed that, after actinomycin treatment: (a) the translational complexity of the mRNA population decreased substantially; (b) at least 66 major mRNAs either increased or decreased significantly in relative abundance; (c) most of the mRNAs which increased in relative abundance were those which, in untreated cells, are most abundant in the short poly(A) fractions; and (d) most of the mRNAs which decreased in relative abundance were those which, in untreated cells, are most abundant in the long poly(A) fractions. These experiments, coupled with our previous results, lead us to suggest that those mRNAs which are enriched in fractions containing short, steady-state size poly(A) tracts are most stable. Our data do not support models in which poly(A) length regulates mRNA stability.Additional experiments in this paper demonstrate that: (a) a previously identified (Palatniket al., 1979) class of mRNA which contains only oligo(A) tracts (approx. 25 nucleotides) is also relatively unstable in the presence of actinomycin D; and (b) a major change in the distribution of translatable mRNAs occurs after cells are removed from prolonged exposure to actinomycin D. Many mRNAs which had been substantially reduced by actinomycin D treatment, increased m relative abundance after removal of the drug. Moreover, a large fraction of the mRNAs which had accumulated to very high levels in actinomycin D-treated cells, decreased dramatically in relative abundance after the drug was removed.