Encalada Lab Research
Intracellular transport is essential for diverse cellular processes including establishment and maintenance of polarity, distribution of organelles, and secretion. Much of this transport is mediated by kinesins and dyneins, two families of molecular motor proteins that track along polarized microtubule paths in plus- and minus-end directed routes, respectively. Transport in axons poses a particular challenge in that it
Regulation of Intracellular Transport and the Role of Axonal Transport in Neurodegeneration
Axonal transport of cargo along axons by kinesin and dynein.
has to be highly processive and regulated to ensure delivery of cargo such as organelles, synaptic vesicle precursors, signaling complexes, etc., to their proper locations, often across very long distances. Yet how motor regulation is accomplished or what the identity of cargo-transport machinery-regulatory complexes within neurons, remains unclear. Our research program is focused on two overarching themes. First, we use a combination of cell biology, genetics, biochemistry, and high-resolution microscopy in mammalian neurons and in the soil nematode Caenorhabditis elegans to identify and characterize cargo-motor complexes, and to characterize the mechanisms of bidirectional motor transport regulation in neurons. Second, we seek to integrate and extend our knowledge of motor regulation to dissect the role that defective vesicular transport plays in the accumulation of misfolded proteins inside axons, and scrutinize the hypothesis that this contributes to or causes neurodegeneration.
Axonal Transport, Protein Spread, and Neurodegeneration: Prions and Tau
Mechanisms of Axonal Transport
Two important questions in motor biology are: (1) what is the identity of all the different cargomotor complexes traveling in neurons? and (2) how is bidirectional transport regulated? We are studying these questions by focusing on identifying the structural and regulatory transport complexes that move vesicles carrying various cargoes including the normal mammalian prion protein (PrP ) and synaptic vesicle precursors in axons, and have identified the motor proteins involved in transport of these vesicles in neurons. We use computational particle tracking and quantitative image analyses, as well as biochemical and genetic approaches, to characterize the dynamics of cargo transport in neurons in mammalian and Caenorhabditis elegans systems.
Model of the mechanism of axonal transport of vesicles carrying the normal mammalian prion protein (PrP ) in neurons (Reprinted from Encalada et al. 2011 Cell).
Our lab is interested in identifying and characterizing signaling and other components that regulate the movement of these vesicles in axons, as well as structural components that bridge cargo to motors. We hope to use our knowledge of these components and mechanisms to build models of axonal transport regulation.
A video of fluorescently-labeled infectious mouse prions in hippocampal neurons.
Recent studies have highlighted the prion-like propagation of proteins involved in a number of neurodegenerative diseases such as Alzheimer’s disease (AD), frontotemporal dementia (FTD), and Parkinson’s disease (PD). Thus, understanding the mechanisms of prion propagation in neurons might unveil the mechanisms of spread of misfolded aggregates in these and other neurodegenerative diseases. We use genetics, high-resolution single-molecule and live microscopy, biochemistry, and molecular biophysical approaches in mammalian neurons, in mice, and in vitro
systems to elucidate the mechanisms of axonal transport and the spread of prions and prion-like proteins including Tau. Our goal is to testthe hypothesis that the spread or prions towards and within the brain is via axonal transport pathways regulated by molecular motors.
Mechanisms of Transthyretin (TTR) Neuronal Toxicity
The transthyretin amyloidoses are a related group of systemic degenerative diseases, wherein secretion of the transthyretin (TTR) tetramer from the liver followed by its dissociation, aberrant misfolding and aggregation causes proteotoxicity and degeneration of post-mitotic tissues including neurons. These diseases include Senile Systemic Amyloidosis (SSA), a highly under-diagnosed but common cardiovascular disease of the elderly, and Familial Amyloid Polyneuropathy (FAP). The peripheral and autonomic nervous systems, as well as the heart are compromised by TTR aggregation in humans, even though these tissues do not synthesize TTR. The mechanistic basis for this cell non-autonomous cytotoxicity as well as the TTR conformations involved, are unknown. Our laboratory has generated C. elegans TTR proteotoxicity models that exhibit TTR aggregation and compelling neuronal phenotypes relevant to human disease, and that point to specific cell non-autonomous targets of TTR toxicity. We use genetic, genomics, biochemistry, high-resolution in vivo live microscopy, and small molecule approaches to characterize the mechanisms of TTR neuronal toxicity. In collaboration with the laboratory of Jeff Kelly (The Scripps Research Institute), our TTR models also provide a platform to elucidate the mechanisms of action of the small molecule drug tafamidis, discovered by the Kelly Lab, that slows rate-limiting TTR tetramer dissociation and amyloidogenesis, and dramatically slows the progression of both the neuropathy and the cardiomyopathy in patients.