Sequences responsible for intracellular localization of beta-actin messenger RNA also affect cell phenotype.
Sequences responsible for intracellular localization of beta-actin messenger RNA also affect cell phenotype.
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DOI：
10.1083/jcb.127.2.441
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发表时间：
1994-10
期刊：
影响因子：
--
通讯作者：
Singer RH
中科院分区：
文献类型：
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作者：
Kislauskis EH;Zhu X;Singer RH
我们通过在鸡胚成纤维细胞（CEFs）中表达LacZ/ β -肌动蛋白嵌合mRNA，确定了将β -胞浆肌动蛋白mRNA直接传递到细胞外周的RNA序列的结构和功能，并将其定位到2段3'-非翻译区。一个54-nt片段，“RNA邮编”，和一个同源但活性较低的43-nt片段，每个定位到前导片的β -半乳糖苷酶活性。该zipcode包含完整的活性，其中的突变或删除会减少，但不会消除其活性，这表明有几个基序对活性有贡献。其中两个基序，当多聚时，可以再生几乎全部的活性。这些序列在进化中是高度保守的，因为人类β -肌动蛋白邮编同样位于3'UTR中，在鸡细胞中也同样定位良好。针对邮政编码的互补硫代寡核苷酸使内源性β -肌动蛋白mRNA脱域，而那些与邮政编码外的区域互补的寡核苷酸或感测寡核苷酸则没有。肌动蛋白mRNA或蛋白水平不受反义处理的影响，但板足结构发生了显著变化，使用相同的反邮政编码寡核苷酸可以观察到肌动蛋白应激纤维的组织。因此，离散的3'UTR序列将β -肌动蛋白同工异构体的合成引导到前导片层，并可能通过肌动蛋白细胞骨架影响细胞形态。
We have characterized the structure and function of RNA sequences that direct beta-cytoplasmic actin mRNA to the cell periphery were mapped to two segments of 3'-untranslated region by expression of LacZ/beta-actin chimeric mRNAs in chicken embryo fibroblasts (CEFs). A 54-nt segment, the "RNA zipcode," and a homologous but less active 43-nt segment each localized beta-galactosidase activity to the leading lamellae. This zipcode contains the full activity, and mutations or deletions within it reduce, but do not eliminate, its activity, indicating that several motifs contribute to the activity. Two of these motifs, when multimerized, can regenerate almost full activity. These sequences are highly conserved in evolution, since the human beta-actin zipcode, positioned identically in the 3'UTR localizes equally well in chicken cells. Complementary phosphorothioate oligonucleotides against the zipcode delocalized endogenous beta-actin mRNA, whereas those complementary to the region just outside the zipcode, or sense oligonucleotides, did not. Actin mRNA or protein levels were unaffected by the antisense treatments, but a dramatic change in lamellipodia structure, and actin stress fiber organization was observed using the same antizipcode oligonucleotides which delocalized the mRNA. Hence, discrete 3'UTR sequences direct beta-actin isoform synthesis to the leading lamellae and affect cell morphology, presumably through the actin cytoskeleton.