Synthesis, transport, and utilization of specific flagellar proteins during flagellar regeneration in Chlamydomonas.
Synthesis, transport, and utilization of specific flagellar proteins during flagellar regeneration in Chlamydomonas.
复制标题
衣原体鞭毛再生期间特定鞭毛蛋白的合成，运输和利用。
DOI：
10.1083/jcb.93.3.615
复制
发表时间：
1982-06
影响因子：
7.8
通讯作者：
Witman, G B
中科院分区：
文献类型：
--
作者：
Remillard, S P;Witman, G B
我们在去鞭毛之前和之后的不同时间用 10 分钟的 35SO4(-2) 脉冲标记衣藻的配子，并在脉冲后立即分离全细胞和鞭毛。通过一维或二维凝胶电泳分离标记的蛋白质，并测定掺入特定蛋白质中的同位素的量。通过将缺失的轴丝多肽与瘫痪突变体的超微结构缺陷相关联，或通过鞭毛部分的多肽分析，鉴定出具有特定结构的单个蛋白质。大多数鞭毛蛋白的合成似乎是在鞭毛截肢后协调诱导的。大多数定量蛋白质的合成率在去鞭毛后增加了至少 4 至 10 倍。结构内包含的蛋白质（例如放射状辐条蛋白质 [RSP]）的合成动力学通常相似；然而，不同结构（例如RSP与α-和β-微管蛋白）所含蛋白质的合成动力学是不同的。大多数新合成的鞭毛蛋白被快速转运到鞭毛中，其动力学反映了细胞器的生长速率；例外情况包括中央小管复合蛋白（CT1）和肌动蛋白样成分，这两种成分似乎几乎完全由预先存在的未标记池提供。同位素稀释实验表明，对于大多数定量的轴丝蛋白来说，用于组装新轴丝的多肽链至少有 35-40% 是在再生过程中合成的；这些蛋白质似乎具有相对于其在轴丝中的化学计量大致相同大小的预去鞭毛池。相比之下，CT1 和肌动蛋白样蛋白的库相对较大。
We labeled gametes of Chlamydomonas with 10-min pulses of 35SO4(-2) before and at various times after deflagellation, and isolated whole cells and flagella immediately after the pulse. The labeled proteins were separated by one- or two-dimensional gel electrophoresis, and the amount of isotope incorporated into specific proteins was determined. Individual proteins were identified with particular structures by correlating missing axonemal polypeptides with ultrastructural defects in paralyzed mutants, or by polypeptide analysis of flagellar fractions. Synthesis of most flagellar proteins appeared to be coordinately induced after flagellar amputation. The rate of synthesis for most quantified proteins increased at least 4- to 10-fold after deflagellation. The kinetics of synthesis of proteins contained together within a structure (e.g., the radial spoke proteins [RSP] ) were frequently similar; however, the kinetics of synthesis of proteins contained in different structures (e.g., RSP vs. alpha- and beta- tubulins) were different. Most newly synthesized flagellar proteins were rapidly transported into the flagellum with kinetics reflecting the rate of growth of the organelle; exceptions included a central tubule complex protein (CT1) and an actinlike component, both of which appeared to be supplied almost entirely from pre-existing, unlabeled pools. Isotope dilution experiments showed that, for most quantified axonemal proteins, a minimum of 35-40% of the polypeptide chains used in assembling a new axoneme was synthesized during regeneration; these proteins appeared to have predeflagellation pools of approximately the same size relative to their stoichiometries in the axoneme. In contrast, CT1 and the actinlike protein had comparatively large pools.