Dutcher Susan K, Morrissette Naomi S, Preble Andrea M, Rackley Craig, Stanga John
Department of Genetics,Washington University School of Medicine, St Louis, Missouri 63110, USA.
Mol Biol Cell. 2002 Nov;13(11):3859-69. doi: 10.1091/mbc.e02-04-0205.
Centrioles and basal bodies are cylinders composed of nine triplet microtubule blades that play essential roles in the centrosome and in flagellar assembly. Chlamydomonas cells with the bld2-1 mutation fail to assemble doublet and triplet microtubules and have defects in cleavage furrow placement and meiosis. Using positional cloning, we have walked 720 kb and identified a 13.2-kb fragment that contains epsilon-tubulin and rescues the Bld2 defects. The bld2-1 allele has a premature stop codon and intragenic revertants replace the stop codon with glutamine, glutamate, or lysine. Polyclonal antibodies to epsilon-tubulin show peripheral labeling of full-length basal bodies and centrioles. Thus, epsilon-tubulin is encoded by the BLD2 allele and epsilon-tubulin plays a role in basal body/centriole morphogenesis.
中心粒和基体是由九个三联体微管片组成的圆柱体,在中心体和鞭毛组装中发挥着重要作用。携带bld2-1突变的衣藻细胞无法组装双联和三联微管,并且在分裂沟定位和减数分裂方面存在缺陷。通过定位克隆,我们已经跨越了720 kb,并鉴定出一个13.2 kb的片段,该片段包含ε-微管蛋白并挽救了Bld2缺陷。bld2-1等位基因有一个过早的终止密码子,基因内回复突变体用谷氨酰胺、谷氨酸或赖氨酸取代了终止密码子。针对ε-微管蛋白的多克隆抗体显示全长基体和中心粒的外周标记。因此,ε-微管蛋白由BLD2等位基因编码,并且ε-微管蛋白在基体/中心粒形态发生中发挥作用。