Rompolas Panteleimon, Pedersen Lotte B, Patel-King Ramila S, King Stephen M
Department of Molecular, Microbial and Structural Biology, University of Connecticut Health Center, Farmington, CT 06030, USA.
J Cell Sci. 2007 Oct 15;120(Pt 20):3653-65. doi: 10.1242/jcs.012773. Epub 2007 Sep 25.
Intraflagellar transport (IFT) is the bi-directional movement of particles along the length of axonemal outer doublet microtubules and is needed for the assembly and maintenance of eukaryotic cilia and flagella. Retrograde IFT requires cytoplasmic dynein 1b, a motor complex whose organization, structural composition and regulation is poorly understood. We have characterized the product of the Chlamydomonas FAP133 gene that encodes a new WD-repeat protein similar to dynein intermediate chains and homologous to the uncharacterized vertebrate protein WD34. FAP133 is located at the peri-basal body region as well as in punctate structures along the flagella. This protein is associated with the IFT machinery because it is specifically depleted from the flagella of cells with defects in anterograde IFT. Fractionation of flagellar matrix proteins indicates that FAP133 associates with both the LC8 dynein light chain and the IFT dynein heavy chain and light intermediate chain (DHC1b-D1bLIC) motor complex. In the absence of DHC1b or D1bLIC, FAP133 fails to localize at the peri-basal body region but, rather, is concentrated in a region of the cytoplasm near the cell center. Furthermore, we found that FAP133, LC8, DHC1b, D1bLIC, the FLA10 kinesin-2 necessary for anterograde IFT and other IFT scaffold components associate to form a large macromolecular assembly.
鞭毛内运输(IFT)是颗粒沿着轴丝外双联微管长度进行的双向运动,是真核生物纤毛和鞭毛组装与维持所必需的。逆行IFT需要细胞质动力蛋白1b,这是一种动力复合体,其组织、结构组成和调控机制尚不清楚。我们对衣藻FAP133基因的产物进行了表征,该基因编码一种新的WD重复蛋白,类似于动力蛋白中间链,与未表征的脊椎动物蛋白WD34同源。FAP133位于基体周围区域以及沿鞭毛的点状结构中。这种蛋白与IFT机制相关,因为它在正向IFT有缺陷的细胞的鞭毛中特异性缺失。鞭毛基质蛋白的分级分离表明,FAP133与LC8动力蛋白轻链以及IFT动力蛋白重链和轻中间链(DHC1b-D1bLIC)动力复合体相关联。在没有DHC1b或D1bLIC的情况下,FAP133无法定位在基体周围区域,而是集中在细胞中心附近的细胞质区域。此外,我们发现FAP133、LC8、DHC1b、D1bLIC、正向IFT所需的FLA10驱动蛋白-2以及其他IFT支架成分相互关联形成一个大型大分子组装体。