The contents of eukaryotic cells are highly dynamic yet organized spatially and temporally. This is achieved primarily by the microtubule cytoskeleton and associated transport machinery, whose fundamental nature is highlighted by the many neurological diseases caused by mutations in them. The overarching goal of the Reck-Peterson lab is to understand how this system works at molecular, cellular, and organismal scales.

The Reck-Peterson lab is highly interdisciplinary, and we use in vitro biochemical reconstitution, protein engineering, single-molecule imaging, proteomics, genoomics, live-cell imaging, and genetics to achieve our goals. Through collaborative projects with others at UC San Diego we use cryo-electron microscopy (with Andres Leschziner’s lab) and cryo-electron tomography (with Elizabeth Villa’s lab) to incorporate a structure-guided approach to understanding intracellular transport, and we develop testable quantitative physical models of transport (with Elena Koslover’s lab).

The Reck-Peterson lab has made major contributions to determining how the dynein motor works and is regulated, to developing tools and screening strategies to study bi-directional movement of cargos on microtubules, to understanding the regulation of intracellular transport in cells, to determine how defects in microtubule-based trafficking may impact Parkinson’s Disease. Current research directions in the lab include:

(1) How does the dynein motor work? Dynein is a large and complicated molecular machine. We have made contributions to developing recombinant systems to express dynein, determining how dynein steps along microtubules, and determining how it is regulated by Lis1. Both the dynein motor encoding gene (DYNC1H1) and the LIS1 gene are mutated in the neurodevelopmental disease lissencephaly. We have shown that the Lis1 protein binds directly to dynein’s motor domain and impacts its mechanochemical cycle (dynein is a AAA ATPase and its nucleotide cycle controls its interactions with its microtubule track). We also discovered that Lis1 promotes the formation of activated dynein complexes. Activated dynein complexes include 12 dynein subunits, 23 dynactin subunits and a dimeric “activating adaptor”. Current efforts in the lab are focused on determining how Lis1 promotes the formation of these massive, activated dynein complexes. For example, in our most recent work in collaboration with the Leschziner lab we captured a structural intermediate in the Lis1-mediated activation of dynein. [bioRxiv]

(2) What is the mechanism and function of organelle hitchhiking? We co-discovered the phenomenon of organelle hitchhiking [pdf]. Using a genetic screen designed to determine how peroxisomes link to molecular motors for movement, we unexpectedly discovered that peroxisomes do not directly recruit dynein and kinesin, but rather hitchhike on motile endosomes. We identified a putative molecular tether (PxdA) and a phosphatase (DipA) that are required for peroxisome hitchhiking. Current efforts are focused on determine how and why peroxisomes hitchhike on early endosomes.

(3) Why do perturbations to the transport machinery cause disease? Mutations in many components of the transport machinery (the motors, the tracks and regulators of both) are directly linked to a wide variety of neurodevelopmental and neurodegenerative diseases. In addition to our studies on Lis1, we recently began working on LRRK2. The LRRK2 gene is one of the most commonly mutated genes in familial Parkinson’s disease. In collaboration with the Leschziner lab, we showed that LRRK2 binds directly to microtubules in the absence of any other proteins. By doing so it can act as a “roadblock” for dynein and kinesin microtubule-based motors, nearly completely abolishing their movement at low nanomolar concentrations. LRRK2 binds to microtubules at lower concentrations when it carries Parkinson’s Disease mutations or in the presence of LRRK2-specific Type 1 kinase inhibitors. Because Type 1 kinase inhibitors are being investigated for the treatment of Parkinson’s Disease, current efforts in the lab are aimed at determining if endogenous LRRK2 interacts with microtubules in the presence of kinase inhibitors.

(4) RNA recoding of molecular motors in cephalopods: Cephalopods such as squid and octopus extensively use Adenosine to Inosine RNA editing to edit their messenger RNAs, which can result in recoding (amino acid changes at the protein level). The transport machinery that we study is extensively recoded. We discovered that this recoding increases in the cold by studying recoding in wild squid. Further, we found that kinesin motors carrying recoded sites were “better” motors in the cold- they were more likely to interact with microtubules when carrying cold-specific recoded sites. We also showed that sites of recoding may serve as a roadmap for identifying regions of proteins that can be functionally modified, suggesting that the cephalopod editomes could serve as a guide for understanding the function of proteins that are conserved between cephalopods and other organisms. Currently, we are developing a database to explore how the squid transcriptome changes in response to temperature based on experiments with wild squid.

(5) Other directions: We are also exploring a number of new directions related to transport in a variety of organisms using a wide range of techniques and methods. If you are interested in joining our team, contact Sam to discuss possible project directions.