Gueth-Hallonet C, Antony C, Aghion J, Santa-Maria A, Lajoie-Mazenc I, Wright M, Maro B
Département de Biologie du Développement, Institut Jacques Monod, CNRS-Université Paris VII, France.
J Cell Sci. 1993 May;105 ( Pt 1):157-66. doi: 10.1242/jcs.105.1.157.
gamma-Tubulin, a recently discovered member of the tubulin superfamily, is a peri-centriolar component considered to be essential for microtubule nucleation. Mouse oocytes and early embryos lack centrioles until the blastocyst stage. Thus, early mouse embryos allowed us to study the location of gamma-tubulin in animal cells in the absence of centrioles. For this, we used an antiserum directed against a specific peptide of the gamma-tubulin sequence, which is conserved among species. This serum has been characterised both in PtK2 and mouse cells. We found that it specifically-stained the spindle poles and the cytoplasmic microtubule organizing centers in metaphase II oocytes and the spindle poles in mitosis during the cleavage stages. In contrast, no interphase staining could be detected during cleavage. Since the overall level of gamma-tubulin did not decrease during interphase, as shown by immunoblotting experiments, this absence of staining during interphase is probably due to a cytoplasmic dispersion of gamma-tubulin. A single dot-like interphase reactivity appeared at the 32-cell stage. In parallel, electron microscopy studies allowed us to detect centrioles for the first time at the 64-cell stage. The possible roles of gamma-tubulin in microtubule nucleation and in centrosome maturation are discussed.
γ-微管蛋白是微管蛋白超家族中最近发现的成员，是一种中心粒周围成分，被认为对微管成核至关重要。小鼠卵母细胞和早期胚胎在囊胚阶段之前缺乏中心粒。因此，早期小鼠胚胎使我们能够在没有中心粒的情况下研究γ-微管蛋白在动物细胞中的定位。为此，我们使用了一种针对γ-微管蛋白序列特定肽段的抗血清，该肽段在物种间是保守的。这种血清已在PtK2细胞和小鼠细胞中进行了鉴定。我们发现，它能特异性地标记中期II卵母细胞中的纺锤体极和细胞质微管组织中心，以及卵裂期有丝分裂过程中的纺锤体极。相比之下，在卵裂过程中未检测到间期染色。免疫印迹实验表明，γ-微管蛋白的总体水平在间期并未降低，因此间期未染色可能是由于γ-微管蛋白在细胞质中分散所致。在32细胞期出现了单个点状的间期反应。同时，电子显微镜研究使我们首次在64细胞期检测到中心粒。本文还讨论了γ-微管蛋白在微管成核和中心体成熟中的可能作用。
