3-D FINE STRUCTURE OF THE C ELEGANS MEIOTIC SPINDLE POLE

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。在减数分裂和有丝分裂过程中，染色体的可靠分离依赖于微管(MT)组装成一个两极纺锤体。线虫卵子的减数分裂纺锤体与受精卵的第一个有丝分裂纺锤体在形态上是不同的，但这两种结构在同一细胞质中形成，间隔约20分钟。减数分裂的MTS不像有丝分裂那样从中心体成核，而是最初围绕着染色质聚合。与染色质相关的正端定向马达蛋白参与了将MT负端推离染色质的过程，然后MT阵列聚焦到两个相对的两极。最近，线虫的mei-1和mei-2基因被发现是减数分裂纺锤体形成所必需的，但不是有丝分裂纺锤体功能所必需的。遗传分析表明，mei-1和mei-2基因是线虫中的katanin同源基因。Katanin是一种异二聚体的MT切断蛋白，由催化亚基和调节亚基组成。功能丧失突变导致无序的减数分裂纺锤体和减数分裂失败，但随后的有丝分裂纺锤体是正常的。相反，功能获得突变导致正常减数分裂，随后在短纺锤体上进行有丝分裂。这些观察表明，Katanin的MT切断活性可能解释了减数分裂纺锤体如何在后来支持更大的有丝分裂纺锤体组装的细胞质环境中保持较小的尺寸。我们制作了两个雌性减数分裂纺锤体的系列断层照片，发现大多数(~80%)的极-近端MT末端是开放的，而在有丝分裂纺锤体的中心体上主要是封闭的/有帽的末端。我们目前正在准备具有mei-1功能获得和功能丧失突变的菌株，以确定在减数分裂过程中，类katanin蛋白的功能如何影响MTS的3D排列和形态。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Faithful chromosome segregation during meiosis and mitosis depends on assembly of the microtubules (MTs) into a bipolar spindle. The meiotic spindle of the C. elegans ovum is morphologically distinct from the first mitotic spindle of the zygote, yet both structures form in the same cytoplasm approximately 20 minutes apart. Instead of nucleating from centrosomes as in mitosis, meiotic MTs polymerize initially around chromatin. Chromatin-associated plus-end directed motor proteins have been implicated in pushing MT minus ends away from the chromatin, and the MT array is then focused into two opposing poles. Recently the mei-1 and mei-2 genes of C. elegans were found to be required for meiotic spindle formation but not for mitotic spindle function. Genetic analysis has shown that mei-1 and mei-2 genes are the katanin orthologues in C. elegans. Katanin is a heterodimeric MT-severing protein consisting of a catalytic and a regulatory subunit. Loss-of-function mutations result in disorganized meiotic spindles and meiotic failure, but the subsequent mitotic spindles are normal. In contrast, gain-of-function mutations result in normal meiosis followed by mitosis on short spindles. These observations suggest that the MT severing activity of katanin may explain how meiotic spindles maintain their small size in a cytoplasm environment that later supports the assembly of a much larger mitotic spindle. We have produced serial tomograms from two female meiotic spindles and found that the majority (~80%) of the pole-proximal MT ends are open, in contrast to the closed/capped ends that predominate at the centrosome of the mitotic spindle. We are currently preparing strains with gain and loss-of-function mutations in mei-1 to determine how the 3D arrangements and morphology of MTs are affected by the function of katanin-like proteins during meiosis.