Regulation of motors in bidirectional motility of early endosomes in the model pathogenic fungus Ustilago maydis

模型病原真菌玉米黑粉菌早期内体双向运动马达的调节

• 批准号: BB/F022956/1
• 负责人: Gero Steinberg
• 金额: $ 44.79万
• 依托单位: UNIVERSITY OF EXETER
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2008
• 资助国家: 英国
• 起止时间: 2008 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FF022956%2F1
• 关键词: Regulation; motors; bidirectional; motility; early

Cell polarization is a fundamental feature of eukaryotic cells and is carried to an extreme in polarized growing cells, such as animal neurons, filamentous fungi, plant pollen tubes and root hairs. In the mammalian axon, expansion of the growth cone is supported by transport of membranous organelles, such as endosomes, synaptic vesicles, proteins and RNA along the fibres of the cytoskeleton, namely microtubules and F-actin. Specialized protein machines, called molecular motors rapidly move along these 'tracks' for delivery of their cargo. Axonal transport happens mainly along microtubules and is essential for brain function and development. Consequently, defects in motor activity result in severe neuro-degenerative diseases. Despite its importance the molecular basis of long-distance transport is not well-understood. This is in part due to the lack of simple and genetically tractable model organism to study long-distance transport. This gap may be filled by filamentous fungi, which are genetically tractable and also grow as highly polarized cells called hyphae. Hyphal growth requires delivery of enzymes, membranes and cell wall-precursors to the expanding tip. Similar to axons, microtubule-based transport is required for tip growth, which involves motor proteins, such as kinesin-3 and kinesin-1, that are also found in mammals. At the growth region enzymes and wall-components are released by a process called exocytosis. Very recently, we have shown in a fungus called Ustilago maydis that endocytosis, which is the uptake of material into the cell, participates in hyphal tip growth and is necessary for the ability of this fungus to cause plant disease. We found that early endosomes (EEs), which are membrane-bound organelles that collect the up-taken material, are of crucial importance. This is most likely achieved by a supportive function of EEs in recycling of enzymes and receptors at the growth region. Interestingly, we also found that EEs rapidly move up and down the hyphae of U. maydis. This is achieved by the two motor proteins, kinesin-3 and dynein, which move endosomes in opposite directions along microtubules. By genetic means we interfered with the balance of their activity, and this resulted in defects in endosomes motility and a block in hyphal elongation. This result strongly implies that the movement itself is essential for fungal tip growth. However, neither the cellular role of endosome motility nor the regulation of the underlying motors is currently known. U. maydis is one of the best established model systems for studying fungal pathogenicity and the role of the cytoskeleton in hyphal growth. U. maydis combines powerful technical advantages, including a published genome, and numerous genetic tools (e.g. inducible promoters) and cytological tools such as GFP, mRFP, CFP, YFP and photoactivatable GFP are established. We will make use of these technical advantages in order to address the following questions: (1) How is bi-directional EE motility regulated und how do motors balance their activity? (2) Which part other of the kinesin-3 motor binds to EEs? (3) What proteins interact with kinesin-3 and which role do these have in EE motility? The project will provide novel insight into the mechanism of hyphal tip growth by fungi. It will therefore be of fundamental interest to all aspects of fungal research, but will particularity stimulate research on fungal pathogenicity. Therefore, our work will be of benefit to the UK pharmaceutical and agricultural biotechnology industries. Of even greater potential significance, however, is that the motor proteins involved (kinesin-3 and dynein) are also important in long-distance axonal transport in neurons. Therefore, the proposed studies promise also to provide a better understanding of motor protein interplay in mammalian cells.

细胞极化是真核细胞的基本特征，并且在极化生长的细胞（例如动物神经元、丝状真菌、植物花粉管和根毛）中被带到极端。在哺乳动物轴突中，生长锥的扩张由膜性细胞器如内体、突触囊泡、蛋白质和RNA沿着细胞骨架的纤维即微管和F-肌动蛋白的运输支持。专门的蛋白质机器，称为分子马达，沿着这些“轨道”快速移动，以运送它们的货物。轴突运输主要沿沿着微管发生，对脑功能和发育至关重要。因此，运动活动的缺陷导致严重的神经退行性疾病。尽管它的重要性，远距离运输的分子基础还没有得到很好的理解。这部分是由于缺乏简单和遗传学上易于处理的模式生物来研究长途运输。这一空白可能被丝状真菌填补，丝状真菌在遗传上是易处理的，并且也以高度极化的细胞（称为菌丝）生长。菌丝生长需要将酶、膜和细胞壁前体输送到扩张的尖端。与轴突类似，尖端生长需要基于微管的运输，这涉及马达蛋白，如驱动蛋白-3和驱动蛋白-1，它们也在哺乳动物中发现。在生长区，酶和细胞壁成分通过一种称为胞吐作用的过程释放出来。最近，我们在一种叫做玉米黑粉菌的真菌中发现，内吞作用（将物质吸收到细胞中）参与菌丝顶端的生长，并且是该真菌引起植物疾病的能力所必需的。我们发现，早期内体（EE），这是膜结合的细胞器，收集的物质，是至关重要的。这很可能是通过EE在生长区域的酶和受体再循环中的支持功能实现的。有趣的是，我们还发现EE在U.玉米粉这是通过两种马达蛋白实现的，驱动蛋白-3和动力蛋白，它们沿着沿着微管以相反的方向移动内体。通过遗传手段，我们干扰了它们活性的平衡，这导致了内体运动的缺陷和菌丝伸长的阻滞。这一结果强烈暗示运动本身对于真菌尖端生长至关重要。然而，无论是内体运动的细胞作用，也不是潜在的电机的调节是目前已知的。联合玉米是研究真菌致病性和细胞骨架在菌丝生长中的作用的最佳建立的模型系统之一。联合maydis结合了强大的技术优势，包括公开的基因组，以及建立了许多遗传工具（例如诱导型启动子）和细胞学工具，例如GFP、mRFP、CFP、YFP和可光活化的GFP。我们将利用这些技术优势，以解决以下问题：（1）双向EE运动是如何调节的，以及如何电机平衡他们的活动？(2)驱动蛋白-3马达的哪个部分与EE结合？(3)哪些蛋白质与驱动蛋白-3相互作用，它们在EE运动中起什么作用？该项目将为真菌菌丝顶端生长的机制提供新的见解。因此，它将是真菌研究的各个方面的根本利益，但将特别刺激真菌致病性的研究。因此，我们的工作将有利于英国制药和农业生物技术产业。然而，更大的潜在意义是，所涉及的运动蛋白（驱动蛋白-3和动力蛋白）在神经元的长距离轴突运输中也很重要。因此，拟议的研究也有望提供一个更好的理解在哺乳动物细胞中的马达蛋白的相互作用。

项目成果

Bidirectional transport and pulsing states in a multi-lane ASEP model

• DOI: 10.1088/1742-5468/2011/09/p09027
• 发表时间: 2011-04
• 期刊: Journal of Statistical Mechanics: Theory and Experiment
• 作者: Congping Lin;G. Steinberg;P. Ashwin

Kinesin-3 and dynein cooperate in long-range retrograde endosome motility along a nonuniform microtubule array.

• DOI: 10.1091/mbc.e11-03-0217
• 发表时间: 2011-10
• 期刊: Molecular biology of the cell
• 影响因子: 3.3
• 作者: Schuster M;Kilaru S;Fink G;Collemare J;Roger Y;Steinberg G
• 通讯作者: Steinberg G

Queueing induced by bidirectional motor motion near the end of a microtubule.

• DOI: 10.1103/physreve.82.051907
• 发表时间: 2010-11
• 期刊: Physical review. E, Statistical, nonlinear, and soft matter physics
• 作者: P. Ashwin;Congping Lin;G. Steinberg

Motor-mediated bidirectional transport along an antipolar microtubule bundle: a mathematical model.

• DOI: 10.1103/physreve.87.052709
• 发表时间: 2013-05
• 期刊: Physical review. E, Statistical, nonlinear, and soft matter physics
• 作者: Congping Lin;P. Ashwin;G. Steinberg

Motors in fungal morphogenesis: cooperation versus competition.

真菌形态发生的动力：合作与竞争。

• DOI: 10.1016/j.mib.2011.09.013
• 发表时间: 2011
• 期刊: Current opinion in microbiology
• 影响因子: 5.4
• 作者: Steinberg G