How Does Phosphorylation Regulate Neurofilament Transport?

这个迫切的项目旨在解决关于磷酸化在神经细丝(NF)运输中的作用的争议。长期以来，神经丝的磷酸化一直被认为可以调节轴突的运输速率，从而为成熟的轴突提供稳定性。3个核因子亚基中最大的(NFH)的C末端延伸的磷酸化被经典地认为减缓了核因子的运输。该项目提出，磷酸化通过促进核因子-核因子结合来抑制运输，导致形成太大的核因子束而无法运输。其他涉及NF亚单位敲除或截断的方法已经提出了磷酸化不调节转运的概念。该项目旨在通过在转基因细胞中表达核因子-H侧臂来解决这一争议。他们将操纵CDK5的活性来改变侧臂的磷酸化，并监测对与核因子束联系的影响。拟议的实验应该提供第一次直接比较侧臂磷酸化的影响，而不会出现损害先前研究的并发症。该项目将为本科生和研究生提供培训和指导，促进他们的出版物，并通过支持他们参加国际会议向广大科学界介绍他们。这项研究还对细胞生物学领域产生了广泛的影响，因为它将继续提供关于所有细胞类型中中间丝“承载”方面的新信息。

This EAGER project aims to address a controversy regarding the role of phosphorylation on neurofilament (NF) transport. Neurofilament phosphorylation has long been considered to regulate their axonal transport rate and in doing so to provide stability to mature axons. Phosphorylation of the C-terminal extension of the largest (NFH) of the 3 NF subunits is classically thought to slow NF transport. This project proposes that phosphorylation inhibits transport by fostering NF-NF associations leading to formation of NF bundles too large to transport. Other approaches involving NF subunit knockout or truncation have advanced the notion that phosphorylation does not regulate transport. This project is set out to address this controversy by expressing the NF-H sidearm in transfected cells. They will manipulate cdk5 activity to alter sidearm phosphorylation and monitor the influence on association with NF bundles. The proposed experiments should provide the first direct comparison of the impact of sidearm phosphorylation without the complications that have compromised prior studies. The project will provide training and mentoring of undergraduate and graduate students, fostering their publications, and introduce them to the scientific community at large by supporting their attendance at international meetings. This research also has broad impacts regarding the field of cell biology, in that it will continue to provide novel information regarding the "load-bearing" aspect of intermediate filaments in all cell types.