Protein Phosphorylation Is a Key Event of Flagellar Disassembly Revealed by Analysis of Flagellar Phosphoproteins during Flagellar Shortening in Chlamydomonas
Protein Phosphorylation Is a Key Event of Flagellar Disassembly Revealed by Analysis of Flagellar Phosphoproteins during Flagellar Shortening in Chlamydomonas
复制标题
通过对衣藻鞭毛缩短过程中鞭毛磷蛋白的分析揭示蛋白质磷酸化是鞭毛分解的关键事件
DOI：
10.1021/pr200428n
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发表时间：
2011-08-01
影响因子：
4.4
通讯作者：
Hippler, Michael
中科院分区：
文献类型：
--
作者：
Pan, Junmin;Naumann-Busch, Bianca;Hippler, Michael
纤毛在细胞分裂之前被分解，这被认为是为了调节适当的细胞周期进程。调节纤毛解体的信号通路尚不清楚。最近的生化和遗传数据表明蛋白质磷酸化在纤毛分解中发挥重要作用。在这里，我们分析了经历缩短的鞭毛的膜/基质部分以及衣藻稳态细胞的鞭毛中的磷蛋白。通过 IMAC 和二氧化钛色谱相结合，采用 IMAC (SIMAC) 顺序洗脱策略，对磷酸肽进行富集，并通过串联质谱法进行分析。从 1296 个磷酸肽光谱计数中总共鉴定出 224 种磷蛋白。已鉴定的磷蛋白包括鞭毛运动蛋白（例如外动力蛋白臂）、鞭毛内转运蛋白以及信号分子（包括蛋白激酶、磷酸酶、G 蛋白和离子通道）。其中 89 种磷蛋白仅在缩短的鞭毛中检测到，而 29 种仅在稳定生长细胞的鞭毛中检测到，表明鞭毛缩短期间蛋白质磷酸化发生了巨大变化。我们的数据表明蛋白质磷酸化是鞭毛分解的关键事件，并为进一步研究蛋白质磷酸化控制的鞭毛组装和分解铺平了道路。
Cilia are disassembled prior to cell division, which is proposed to regulate proper cell cycle progression. The signaling pathways that regulate cilia disassembly are not well-understood. Recent biochemical and genetic data demonstrate that protein phosphorylation plays important roles in cilia disassembly. Here, we analyzed the phosphoproteins in the membrane/matrix fraction of flagella undergoing shortening as well as flagella from steady state cells of Chlamydomonas. The phosphopeptides were enriched by a combination of IMAC and titanium dioxide chromatography with a strategy of sequential elution from IMAC (SIMAC) and analyzed by tandem mass spectrometry. A total of 224 phosphoproteins derived from 1296 spectral counts of phosphopeptides were identified. Among the identified phosphoproteins are flagellar motility proteins such as outer dynein arm, intraflagellar transport proteins as well as signaling molecules including protein kinases, phosphatases, G proteins, and ion channels. Eighty-nine of these phosphoproteins were only detected in shortening flagella, whereas 29 were solely in flagella of steady growing cells, indicating dramatic changes of protein phosphorylation during flagellar shortening. Our data indicates that protein phosphorylation is a key event in flagellar disassembly, and paves the way for further study of flagellar assembly and disassembly controlled by protein phosphorylation.