Rev-dependent association of the intron-containing HIV-1 gag mRNA with the nuclear actin bundles and the inhibition of its nucleocytoplasmic transport by latrunculin-B
Rev-dependent association of the intron-containing HIV-1 gag mRNA with the nuclear actin bundles and the inhibition of its nucleocytoplasmic transport by latrunculin-B
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DOI：
10.1046/j.1365-2443.2000.00326.x
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发表时间：
2000-04-01
期刊：
影响因子：
2.1
通讯作者：
Fujisawa, J
中科院分区：
文献类型：
--
作者：
Kimura, T;Hashimoto, I;Fujisawa, J
背景：HIV-1基因表达的一个标志是未剪接的基因组RNA，它也作为Gag/Pol表达的mRNA，输出到细胞质中。雷夫通过核出口信号（NES）指挥这种运输。结果：荧光原位杂交和免疫细胞化学表明，gag mRNA， Rev及其NES受体，CRM1和RanGTPase形成了与潜在的β -肌动蛋白束一致的核轨道。电镜下证实肌动蛋白束形成。这些束是在含有rev的gag RNP形成时观察到的。束的丢失与gag mRNA的核保留有关。细胞质和细胞核gag mRNA的逆转录-聚合酶链反应分析表明，F-actin解聚化合物latrunculin-B （latb）破坏核肌动蛋白丝的形成，导致gag mRNA的输出受到剂量依赖性抑制。mrna阳性细胞的差异亚型从形态学上证实了latb治疗的效果。该化合物对gag mRNA的输出抑制作用是特异性的，而对完全剪接的HIV-1 mRNA和细胞GAPDH mRNA的输出不受该化合物的影响。结论：核β -肌动蛋白束可能在功能上参与了含内含子的HIV-1 gag mRNA的rev依赖性核胞质转运。
Background: A hallmark of HIV-1 gene expression is that unspliced genomic RNA, which also acts as mRNA for the expression of Gag/Pol, is exported to the cytoplasm. Rev directs this transport through the nuclear export signal (NES).Results: Fluorescence in situ hybridization and immunocytochemistry demonstrated that gag mRNA, Rev, and its NES receptor, CRM1, and RanGTPase formed nuclear tracks which were congruent with underlying beta-actin bundles. Actin bundle formation was confirmed electron-microscopically. These bundles were observed upon Rev-containing gag RNP formation. The loss of bundles was associated with the nuclear retention of gag mRNA. Reverse transcription-polymerase chain reaction analysis of both cytoplasmic and nuclear gag mRNAs demonstrated that disruption of nuclear actin filament formation by latrunculin-B (LAT-B), an F-actin depolymerizing compound, resulted in the dose-dependent inhibition of gag mRNA export. The differential subtyping of the mRNA-positive cells confirmed morphologically the effect of LAT-B treatment. The export inhibition was specific to gag mRNA and export of fully spliced HIV-1 tat/rev mRNAs as well as cellular GAPDH mRNA was not affected by the compound.Conclusions: Nuclear beta-actin bundles are suggested to be functionally involved in the Rev-dependent nucleocytoplasmic transport of intron-containing HIV-1 gag mRNA.