Targeted gene disruption of dynein heavy chain 7 of Tetrahymena thermophila results in altered ciliary waveform and reduced swim speed
Targeted gene disruption of dynein heavy chain 7 of Tetrahymena thermophila results in altered ciliary waveform and reduced swim speed
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DOI：
10.1242/jcs.007369
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发表时间：
2007-09-01
影响因子：
4
通讯作者：
Hennessey, Todd M.
中科院分区：
文献类型：
--
作者：
Wood, Christopher R.;Hard, Robert;Hennessey, Todd M.
嗜热四膜虫通过覆盖在其身体上的数百根纤毛的协调跳动来游泳。有人提出纤毛轴丝的外臂动力蛋白控制拍频，而内臂动力蛋白控制波形。为了测试这些内臂之一，动力蛋白重链7蛋白（Dyh7p）的作用，通过营养大核的靶向生物射弹转化产生敲除突变体。通过基因组和cDNA模板的PCR检测证实了编码Dyh7p的基因DYH7的破坏。无论是完整的和洗涤剂提取，这些突变体，我们称之为DYH7neo3，重新激活的细胞模型制剂，显示游泳速度几乎是野生型细胞的一半。虽然DYH7neo3突变体比野生型慢，但它们能够调节它们的游泳速度，并显示对去极化刺激的纤毛逆转。高速视频显微镜的完整的，自由游泳DYH7neo3突变体揭示了不规则的模式纤毛节拍和波形。突变体纤毛似乎参与不太协调的旋转运动，其中在野生型纤毛中看到的典型形状，周期性和协调性缺失或受到干扰。我们建议，轴丝内臂动力蛋白重链7蛋白有助于正常纤毛波形的形成，这反过来又决定了这些细胞的向前游泳速度。
Tetrahymena thermophila swims by the coordinated beating of hundreds of cilia that cover its body. It has been proposed that the outer arm dyneins of the ciliary axoneme control beat frequency, whereas the inner arm dyneins control waveform. To test the role of one of these inner arms, dynein heavy chain 7 protein ( Dyh7p), a knockout mutant was generated by targeted biolistic transformation of the vegetative macronucleus. Disruption of DYH7, the gene which encodes Dyh7p, was confirmed by PCR examination of both genomic and cDNA templates. Both intact and detergent extracted, reactivated cell model preparations of these mutants, which we call DYH7neo3, displayed swim speeds that were almost half that of wildtype cells. Although the DYH7neo3 mutants were slower than wild type, they were able to modulate their swim speed and show ciliary reversal in response to depolarizing stimuli. High-speed video microscopy of intact, free-swimming DYH7neo3 mutants revealed an irregular pattern of ciliary beat and waveform. The mutant cilia appeared to be engaging in less coordinated, swiveling movements in which the typical shape, periodicity and coordination seen in wild-type cilia were absent or disturbed. We propose that the axonemal inner arm dynein heavy chain 7 proteins contribute to the formation of normal ciliary waveform, which in turn governs the forward swimming velocity of these cells.