Ikeda Kazuho, Brown Jennifer A, Yagi Toshiki, Norrander Jan M, Hirono Masafumi, Eccleston Eric, Kamiya Ritsu, Linck Richard W
Department of Biological Sciences, Graduate School of Science, University of Tokyo, Tokyo 113-0033, Japan.
J Biol Chem. 2003 Feb 28;278(9):7725-34. doi: 10.1074/jbc.M210751200. Epub 2002 Nov 14.
Ciliary and flagellar axonemes are basically composed of nine outer doublet microtubules and several functional components, e.g. dynein arms, radial spokes, and interdoublet links. Each A-tubule of the doublet contains a specialized "ribbon" of three protofilaments composed of tubulin and other proteins postulated to specify the three-dimensional arrangement of the various axonemal components. The interdoublet links hold the doublet microtubules together and limit their sliding during the flagellar beat. In this study on Chlamydomonas reinhardtii, we cloned a cDNA encoding a 71,985-Da polypeptide with three DM10 repeats, two C-terminal EF-hand motifs, and homologs extending to humans. This polypeptide, designated as Rib72, is a novel component of the ribbon compartment of flagellar microtubules. It remained associated with 9-fold arrays of doublet tubules following extraction under high and low ionic conditions, and anti-Rib72 antibodies revealed an approximately 96-nm periodicity along axonemes, consistent with Rib72 associating with interdoublet links. Following proteolysis- and ATP-dependent disintegration of axonemes, the rate of cleavage of Rib72 correlated closely with the rate of sliding disintegration. These observations identify a ribbon-associated protein that may function in the structural assembly of the axoneme and in the mechanism and regulation of ciliary and flagellar motility.
纤毛和鞭毛轴丝基本上由九对外周双联微管和几种功能成分组成,例如动力蛋白臂、辐条和双联微管间连接。双联微管的每根A微管都包含由微管蛋白和其他蛋白质组成的三条原纤维的特殊“带”,推测这些蛋白质可确定各种轴丝成分的三维排列。双联微管间连接将双联微管连接在一起,并在鞭毛摆动期间限制它们的滑动。在这项对莱茵衣藻的研究中,我们克隆了一个cDNA,它编码一种71,985道尔顿的多肽,该多肽具有三个DM10重复序列、两个C端EF手基序,并且其同源物延伸至人类。这种多肽被命名为Rib72,是鞭毛微管带区的一种新成分。在高离子强度和低离子强度条件下提取后,它仍与九倍排列的双联微管结合,抗Rib72抗体显示沿轴丝有大约96纳米的周期性,这与Rib72与双联微管间连接结合一致。在轴丝经蛋白酶解和ATP依赖的解体后,Rib72的裂解速率与滑动解体速率密切相关。这些观察结果鉴定出一种与带相关的蛋白质,它可能在轴丝的结构组装以及纤毛和鞭毛运动的机制与调节中发挥作用。