Molecular basis for dyneinopathies reveals insight into dynein regulation and dysfunction
Abstract
Cytoplasmic dynein plays critical roles within the developing and mature nervous systems, including effecting nuclear migration, and retrograde transport of various cargos. Unsurprisingly, mutations in dynein are causative of various developmental neuropathies and motor neuron diseases. These 'dyneinopathies' define a broad spectrum of diseases with no known correlation between mutation identity and disease state. To circumvent complications associated with dynein studies in human cells, we employed budding yeast as a screening platform to characterize the motility properties of seventeen disease-correlated dynein mutants. Using this system, we determined the molecular basis for several classes of etiologically related diseases. Moreover, by engineering compensatory mutations, we alleviated the mutant phenotypes in two of these cases, one of which we confirmed with recombinant human dynein. In addition to revealing molecular insight into dynein regulation, our data provide additional evidence that the type of disease may in fact be dictated by the degree of dynein dysfunction.
Data availability
All of the data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for all figures.
Article and author information
Author details
Funding
Muscular Dystrophy Association (376387)
- Matthew G Marzo
- Jacqueline M Griswold
- Kristina M Ruff
- Rachel E Buchmeier
- Steven M Markus
National Institute of General Medical Sciences (GM 118492)
- Matthew G Marzo
- Steven M Markus
National Institute of General Medical Sciences (GM 112893)
- Colby P Fees
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2019, Marzo et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Download links
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Further reading
-
- Cell Biology
Understanding how the brain controls nutrient storage is pivotal. Transient receptor potential (TRP) channels are conserved from insects to humans. They serve in detecting environmental shifts and in acting as internal sensors. Previously, we demonstrated the role of TRPγ in nutrient-sensing behavior (Dhakal et al., 2022). Here, we found that a TRPγ mutant exhibited in Drosophila melanogaster is required for maintaining normal lipid and protein levels. In animals, lipogenesis and lipolysis control lipid levels in response to food availability. Lipids are mostly stored as triacylglycerol in the fat bodies (FBs) of D. melanogaster. Interestingly, trpγ deficient mutants exhibited elevated TAG levels and our genetic data indicated that Dh44 neurons are indispensable for normal lipid storage but not protein storage. The trpγ mutants also exhibited reduced starvation resistance, which was attributed to insufficient lipolysis in the FBs. This could be mitigated by administering lipase or metformin orally, indicating a potential treatment pathway. Gene expression analysis indicated that trpγ knockout downregulated brummer, a key lipolytic gene, resulting in chronic lipolytic deficits in the gut and other fat tissues. The study also highlighted the role of specific proteins, including neuropeptide DH44 and its receptor DH44R2 in lipid regulation. Our findings provide insight into the broader question of how the brain and gut regulate nutrient storage.
-
- Cell Biology
- Immunology and Inflammation
The endothelial blood-brain barrier (BBB) strictly controls immune cell trafficking into the central nervous system (CNS). In neuroinflammatory diseases such as multiple sclerosis, this tight control is, however, disturbed, leading to immune cell infiltration into the CNS. The development of in vitro models of the BBB combined with microfluidic devices has advanced our understanding of the cellular and molecular mechanisms mediating the multistep T-cell extravasation across the BBB. A major bottleneck of these in vitro studies is the absence of a robust and automated pipeline suitable for analyzing and quantifying the sequential interaction steps of different immune cell subsets with the BBB under physiological flow in vitro. Here, we present the under-flow migration tracker (UFMTrack) framework for studying immune cell interactions with endothelial monolayers under physiological flow. We then showcase a pipeline built based on it to study the entire multistep extravasation cascade of immune cells across brain microvascular endothelial cells under physiological flow in vitro. UFMTrack achieves 90% track reconstruction efficiency and allows for scaling due to the reduction of the analysis cost and by eliminating experimenter bias. This allowed for an in-depth analysis of all behavioral regimes involved in the multistep immune cell extravasation cascade. The study summarizes how UFMTrack can be employed to delineate the interactions of CD4+ and CD8+ T cells with the BBB under physiological flow. We also demonstrate its applicability to the other BBB models, showcasing broader applicability of the developed framework to a range of immune cell-endothelial monolayer interaction studies. The UFMTrack framework along with the generated datasets is publicly available in the corresponding repositories.