Intraflagellar transport proteins are involved in thrombocyte filopodia formation and secretion.
Intraflagellar transport proteins are involved in thrombocyte filopodia formation and secretion.
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DOI：
10.1080/09537104.2017.1361524
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发表时间：
2018-12
期刊：
影响因子：
3.3
通讯作者：
Jagadeeswaran P
中科院分区：
文献类型：
--
作者：
Radhakrishnan U;Alsrhani A;Sundaramoorthi H;Khandekar G;Kashyap M;Fuchs JL;Perkins BD;Omori Y;Jagadeeswaran P
鞭毛内转运（IFT）蛋白对纤毛的发生和维持至关重要。我们在缺乏初级纤毛的斑马鱼血小板中鉴定出ift 122转录本是出乎意料的。IFT蛋白质在纤毛中起运输作用，纤毛的狭窄尺寸可能需要IFT从祖先真核生物中的囊泡运输进化而来。我们假设，IFTs也可能促进运输内的丝状伪足时，形成血小板被激活。为了测试这种可能性，我们敲低ift 122的表达注入反义吗啉代寡核苷酸（MO）到斑马鱼胚胎。激光诱导的动脉血栓形成显示血管闭塞时间（TTO）延长，这与血小板功能缺陷的预期一致。成年斑马鱼中的急性效应通过ift 122的Vivo-Morpholino（Vivo-MO）敲低来评估。体内-MO morphants表现出延长的时间血小板聚集（TTA）在平板倾斜试验后，血小板活化以下激动剂：ADP，胶原蛋白，PAR 1肽，肾上腺素。ATP的发光测定显示，血小板分泌ATP减少在胶原活化的血液中的Vivo-MO ift 122 morphants。此外，DiI-C18标记的变形血小板暴露于胶原蛋白显示丝状伪足的数量和长度减少。在我们的动脉激光血栓形成试验中，对其中已注意到纤毛缺陷的IFT突变体的分析也显示TTO延长。此外，野生型血小板的胶原活化导致丝状伪足内和基部的IFFT 122浓度。总之，这些结果表明，IFT蛋白参与丝状伪足的延伸和ATP的分泌，这是血小板功能的关键。
Intraflagellar transport (IFT) proteins are vital for the genesis and maintenance of cilia. Our identification of ift122 transcripts in zebrafish thrombocytes that lack primary cilia was unexpected. IFT proteins serve transport in cilia, whose narrow dimensions may have necessitated the evolution of IFT from vesicular transport in ancestral eukaryotes. We hypothesized that IFTs might also facilitate transport within the filopodia that form when thrombocytes are activated. To test this possibility, we knocked down ift122 expression by injecting antisense Morpholino oligonucleotides (MOs) into zebrafish embryos. Laser-induced arterial thrombosis showed prolonged time to occlusion (TTO) of the vessel, as would be expected with defective thrombocyte function. Acute effects in adult zebrafish were evaluated by Vivo-Morpholino (Vivo-MO) knockdown of ift122. Vivo-MO morphants showed a prolonged time to thrombocyte aggregation (TTA) in the plate tilt assay after thrombocyte activation by the following agonists: ADP, collagen, PAR1 peptide, and epinephrine. A luminescence assay for ATP revealed that ATP secretion by thrombocytes was reduced in collagen-activated blood of Vivo-MO ift122 morphants. Moreover, DiI-C18 labeled morphant thrombocytes exposed to collagen showed reductions in filopodia number and length. Analysis of ift mutants, in which cilia defects have been noted, also showed prolongation of TTO in our arterial laser thrombosis assay. Additionally, collagen activation of wild-type thrombocytes led to a concentration of IFT122 both within and at the base of filopodia. Taken together these results suggest that IFT proteins are involved in both the extension of filopodia and secretion of ATP, which are critical in thrombocyte function.
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