Analysis of microtubule-organizing centers (MTOCs) in Aspergillus nidulans and Neurospora crassa

真核细胞中的微管形成依赖于称为微管组织中心（MTOCs）的大型多亚基蛋白质复合物。其中一个被研究得非常充分的MTOC是酿酒酵母中的MTOC，其中MTOC嵌入到核膜中并被命名为纺锤极体（SPB）。在裂糖菌pombe中，三种不同的MTOCs在细胞周期的不同阶段产生MT阵列，位于核膜的多个间期iMTOCs， spb和赤道eMTOCs，在细胞分裂期间产生MT阵列。这三者在空间和时间上都是受调控的。在发现γ -微管蛋白的丝状真菌中，描述了两种稳定的MTOCs， spb和间隔MTOCs （sMTOCs）。我们发现sMTOCs通过紊乱蛋白Spa10锚定在间隔上，并且蛋白质组成类似于SPBs的外斑块之一。在粗神经孢子虫中，SPBs聚合有丝分裂和细胞质的MTs，但在细胞质的生长尖端也有MTOCs的证据。对其生化成分和活性一无所知。在此，我们提出了一种比较研究的方法，对针叶草和草的MTOCs进行比较研究。将研究几种蛋白质的作用，如MOZART 1 （MztA和MTZ-1）或含有j结构域的蛋白RspA。除了MTOCs的生化组成外，组装的层次结构将被解开。此外，将采用非靶向方法，如BioID或GFP-trap技术，以发现新的结构和调节成分。

Microtubule formation in eukaryotic cells depends on large, multi-subunit protein complexes, called Microtubule-organizing centres (MTOCs). One extremely well-studied MTOC is the one in Saccharomyces cerevisiae, where the MTOC is embedded into the nuclear envelope and named spindle-pole body (SPB). In Schizosaccharomyces pombe three different MTOCs generate the MT arrays during different stages of the cell cycle, multiple interphase iMTOCs located in the nuclear envelope, the SPBs, and equatorial, eMTOCs, during cyokinesis. All three are spatially and temporally regulated. In the filamentous fungus Aspergillus nidulans, in which gamma-tubulin was discovered, two stable MTOCs were described, the SPBs and septal MTOCs (sMTOCs). We found that sMTOCs are anchored at septa through the disordered protein Spa10, and that the protein composition resembles the one of the outer plaque of SPBs. In Neurospora crassa SPBs polymerize mitotic and cytoplasmic MTs, but in addition there is evidence for MTOCs in the growing tip in the cytoplasm. Nothing is known about the biochemical composition and the activity. Here, we propose to perform a comparative study of the MTOCs in A. nidulans and N. crassa. The role of several proteins, such as MOZART 1 (MztA and MTZ-1) or the J-domain containing protein RspA, will be studied. Besides the biochemical composition of MTOCs, the hierarchy of the assembly will be unraveled. In addition, non-targeted approaches, such as BioID or GFP-trap technology, will be undertaken to discover novel structural and regulatory components.