Richard Vallee, PhD
My lab is interested in a variety of biological phenomena involving motor proteins, with a major emphasis on cytoplasmic dynein. We described it originally as the motor for retrograde axonal transport, but it is now known to have important functions in mitosis, cell migration, growth cone motility, virus transport, and other aspects of neuronal and nonneuronal cell behavior, many of which are under investigation in the lab.
One project involves the role of cytoplasmic dynein in the human brain developmental disease lissencephaly and in normal development. Lissencephaly (smooth brain) arises from mutations in the dynein regulator, LIS1. Investigation of the role of LIS1 in developing rat brain involves transfection of neural progenitor cells by in utero electroporation. We use shRNAs to interfere with expression of LIS1, cytoplasmic dynein, and a variety of additional dynein regulators, including NudE, NudEL, BicD2, and CENP-F. We coexpress fluorescent fusion protein markers to monitor cellular and subcellular behavior with the goal of understanding the specific roles of these factors in the mechanism of neuronal migration, morphogenesis, and proliferation. These studies have led to a model for how LIS1 mutations cause lissencephaly (Tsai et al., 2005; Tsai et al., 2007). They have also led to models for how the forces generated by cytoplasmic dynein in developing neurons contribute to neural progenitor cell migration and division.
A more recent focus of the lab has been on the earliest stages of neurogenesis in the CNS, addressing the unusual behavior of the radial glial progenitor (RGP) cells. These highly elongated cells span the developing neocortex from the ventricles to the brain surface, and serve as guides for migrating neurons. In addition, RGP cells multiply to give rise to most neurons and glial cells in the developing brain, and also to adult neural stem cells. The RGP cells exhibit a remarkable form of cell cycle-dependent nuclear oscillation. Nuclei divide at the surface of the ventricle, ascend basally during G1, undergo S-phase, and then return to the ventricle during G2 to divide again. The physiological purpose and underlying mechanism for this highly conserved behavior have remained largely unexplored until recently. The lab has found that kinesin-3 is responsible for basal nuclear migration in rat brain, and cytoplasmic dynein for apical migration (Tsai et al., 2010). Recent work has revealed that nuclear pores in RGP cells recruit cytoplasmic dynein using two G2-specific mechanisms (Hu et al., 2013). How this mechanism is triggered in early G2, and how mitosis is delayed until nuclei contact the ventricular surface, remain the focus of additional studies.
The lab has also investigated the molecular mechanisms by which LIS1 and other dynein regulators control dynein activity. A number of factors regulate dynein targeting to subcellular sites. We have found LIS1, aided by NudE, to bind to the dynein motor domain during its power stroke, and to stabilize the interaction of dynein with microtubules during this phase of the crossbridge cycle (McKenney et al., 2010; with S. Gross, UC Irvine). The result is a substantial increase in total forces generated by groups of dynein molecules, e. g., those associated with nuclei in the developing brain. We are also investigating the mechanism responsible for dynactin regulation of dynein motor activity, which has led to the identification of a novel processivity-controlling domain, which is subject to autoinhibitory regulation. We are also investigating an emerging class of “RDD” proteins involved in Rab-mediated dynein/dynactin regulation of cargo transport. Additional studies involve the characterization of motor defects in human neurodegenerative mutant forms of cytoplasmic dynein (with C. Fiorello). We have also identified a force-regulating domain within recombinant dynein motor domain (with A. Gennerich, AECOM), which has important implications for understanding the complex intramolecular, as well as intermolecular mechanisms involved in dynein motor regulation.
As a complement to these studies, we continue to investigate mechanisms of dynein regulation in a variety of cellular processes, including vesicular transport in axons and nonneuronal cells, growth cone and lamellipodial formation and function, axonogenesis in cell culture and in live brain tissue, and mitosis. We are also investigating mechanisms used by a non-physiological form of dynein cargo, adenovirus, to hijack dynein and kinesins for use in transport within the infected cell. We have identified specific motor protein subunits recruited to adenovirus, and the capsid components which serve as receptors (Bremner et al., 2009). We recently found that PKA activation during adenovirus infection results in phosphorylation of the dynein LIC1 subunit. The result is displacement of dynein from physiological organelles for recruitment by the virus. We are using high temporal and spatial resolution particle tracking analysis (Yi et al., 2011) to monitor the behavior of fluorescently tagged virus in infected cells. These studies are aimed at determining the role of microtubule plus and minus end-directed motors in the overall infection process. Our data suggest that motor proteins recruited to the virus could facilitate infection, or, alternatively, act in defense of the cell. Our data so far point to at least one new motor-protein based host defense mechanism, an understanding of which may be of general importance in controlling host-pathogen competition.
Carabalona, A., Hu, D. J., and Vallee, R. B (2016) KIF1A Inhibition Immortalizes Brain Stem Cells but Blocks BDNF-mediated Neuronal Migration. Nature Neurosci. 19:253-262.
Reddy,B.,Narayanareddy, J., Mattson-Hoss, M., Wynne, C., Vadpey, O., Durra, A., Chapman, D., Vallee, R. B., and Gross, S. (2016) Load induced Enhancement of Dynein Force Production by LIS1-NudE in Vivo and in Vitro. Nature Commun. 7:12259.
Doobin, D. J.*, Dantas, T.*, Kemal, S.*, and Vallee, (2016) R. B. Nde1-mediated Microcephaly Results from Interference with Neural Progenitor Cell Cycle Progression at Multiple Stages. Nature Commun., 7:12551.
Scherer, J., and Vallee, R. B. (2015) Conformational Changes in Adenovirus Hexon Subunit Responsible for Regulating Cytoplasmic Dynein Recruitment. J. Virol. 89:1013-23.
Nicholas, M. P.,* Hook, P.,* Brenner, S., Lazar, C., Vallee, R. B., + and Gennerich, A.+ (2015) Control of cytoplasmic dynein force production and processivity by its C-terminal domain Nature Commun. 6:6206.
Baffet, A. D., Hu, D. J., and Vallee, R. B. (2013) Cdk1 Activates Pre-Mitotic Nuclear Envelope Dynein Recruitment and Apical Nuclear Migration in Neural Stem cells. Dev. Cell 6:703-716.
Taylor, S. P,* Dantas, T. J.,* Duran, I., Wu, S., Lachman, R., Nelson, S., Cohn, D., Vallee, R. B., and Krakow, D. (2015) Mutations in DYNC2LI1 disrupt cilia function and cause short rib polydactyly syndrome. Nature Commun. 6:7092
Yi., J., Khobrekar, N. V, Dantas, T. J., Zhou, J., and Vallee, R. B. (2016) Imaging of motor-dependent transport in neuronal and nonneuronal cells at high spatial and temporal resolution Meth. Cell Biol. 131:453-65
Baffet, A., D., Carabalona, A., Dantas, T. J., Doobin, D., Hu, D., and Vallee, R. B. (2016) Cellular and Subcellular Imaging of Motor Protein-based Behavior in Embryonic Rat Brain Meth. Cell Biol. 131:349