Microtubule-based intracellular transport is critical for the function of all eukaryotic cells. Cytoplasmic dynein is a minus-end-directed microtubule motor that transports a variety of cargoes, and dynein-cargo interactions require the dynactin complex and specific cargo adapters. How dynein-cargo interactions are regulated in vivo and how dynein activity is coordinated with cargo binding are major open questions in the field. Recently, structural studies and in vitro motility assays led to the hypothesis that the binding of cargo to the pointed end of dynactin's Arp1 filament may unlock dynactin's p150 subunit via a change in its conformation, leading to dynein activation. However, in vivo evidence supporting this hypothesis is missing and the identity of proteins involved in p150's conformational change remain to be defined. We previously found that dynactin's pointed-end protein p25 in Aspergillus nidulans is important for dynein- early endosome interaction. Recently, we discovered that p25 is required for the interaction between dynein-dynactin and HookA, the early endosome dynein adapter. Based on our recent data, we hypothesize that p25 is a key protein regulating dynactin conformation.
Specific Aim 1 is to reveal the roles of dynactin p25 and the cargo adapter HookA in regulating dynactin. Sensitized emission-based FRET imaging, microtubule pelleting and in vitro motility assays will be used to achieve this aim. In addition, our recent genetic work has uncovered a vezatin-like protein, VezA, as a novel regulator for dynein-cargo interaction. VezA is not a cargo adapter like HookA but affects the accumulation of dynein at the cargo-loading site, the microtubule plus end. How VezA regulates dynein remains a mystery.
Specific Aim 2 is to determine the mechanism of VezA action in dynein-mediated vesicle transport. We will use proteomic and genetic approaches to identify new players in the VezA-dynein pathway.

Public Health Relevance

This project is focused on the in vivo regulation of cytoplasmic dynein, a motor that drives many cellular cargoes. The proper function of cytoplasmic dynein is crucial for human health as mutations in dynein or its regulators, such as dynactin and Lis1, are causally linked to brain developmental disorders such as lissencephaly and devastating neurodegenerative diseases such as Amyotrophic lateral sclerosis (ALS or 'Lou Gehrig's Disease).

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM121850-03
Application #
9670833
Study Section
Nuclear and Cytoplasmic Structure/Function and Dynamics Study Section (NCSD)
Program Officer
Ainsztein, Alexandra M
Project Start
2017-05-01
Project End
2021-03-31
Budget Start
2019-04-01
Budget End
2020-03-31
Support Year
3
Fiscal Year
2019
Total Cost
Indirect Cost
Name
Henry M. Jackson Fdn for the Adv Mil/Med
Department
Type
DUNS #
144676566
City
Bethesda
State
MD
Country
United States
Zip Code
20817
Qiu, Rongde; Zhang, Jun; Xiang, Xin (2018) p25 of the dynactin complex plays a dual role in cargo binding and dynactin regulation. J Biol Chem 293:15606-15619
Zhang, Jun; Qiu, Rongde; Xiang, Xin (2018) The actin capping protein in Aspergillus nidulans enhances dynein function without significantly affecting Arp1 filament assembly. Sci Rep 8:11419
Xiang, Xin (2018) Nuclear movement in fungi. Semin Cell Dev Biol 82:3-16