Targeted disruption of an essential RNA-binding protein perturbs cell division in Trypanosoma brucei.
Targeted disruption of an essential RNA-binding protein perturbs cell division in Trypanosoma brucei.
复制标题
有针对性地破坏一种重要的 RNA 结合蛋白会扰乱布氏锥虫的细胞分裂。
DOI：
10.1016/s0166-6851(01)00322-x
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发表时间：
2001
影响因子：
1.5
通讯作者：
Boothroyd,JC
中科院分区：
文献类型：
--
作者：
Manger,ID;Boothroyd,JC
为了进一步研究 RRM1（一种含有许多共有核酸结合基序的蛋白质）的功能，通过同源重组对该蛋白质进行了靶向破坏。为了促进这一点，使用四环素(tet)抑制型启动子来驱动RRM1开放阅读框的辅助拷贝。提供从替代基因座转录的RRM1使得能够选择RRM1基因的2个天然等位基因被破坏的布氏锥虫。以这种方式获得的双敲除细胞在tet存在下生长。从培养基中去除tet会抑制辅助基因的转录和RRM1基因产物的合成，导致高度伸长的多核细胞和具有单一动质体的无核细胞的积累。这些形态学变化发生在 RRM1 缺陷细胞中检测到任何反式剪接或蛋白质合成缺陷之前。这些结果证明 RRM1 在锥虫 RNA 加工中具有特定的作用，而不是更普遍的作用。
To further investigate the function of RRM1 (a protein containing a number of consensus nucleic acid binding motifs), targeted disruption of this protein was undertaken by homologous recombination. To facilitate this, a tetracycline (tet)-repressible promoter was used to drive an auxiliary copy of theRRM1open reading frame. Provision of of RRM1 transcribed from an alternative locus enabled the selection ofTrypanosoma bruceiin which the 2 natural alleles of theRRM1gene were disrupted. The double-knockout cells obtained in this way grew in the presence of tet. Removal of tet from the medium repressed transcription of the auxiliary gene and synthesis of theRRM1gene product, causing the accumulation of highly elongated, multinucleated cells and anucleate cells with a single kinetoplast. These morphological changes occurred before any defect intrans-splicing or protein synthesis could be detected in the RRM1-deficient cells. These results argue for a specific rather than a more general role for RRM1 in trypanosome RNA processing.