The Molecular NeuroPathobiology Laboratory
Marta Budzinska: Mechanisms controlling the axonal retrograde transport pathway and neurotrophic signaling in motor neurons
Axonal transport is the main process enabling fast delivery of organelles and biological molecules inside neurons. Neurotrophin (NT) signalling is of particular importance for large cells such as motor neurons. NTs bind to their receptors at the synapse and, internalised, are transported to the soma where they elicit transcriptional response. This long-range signalling is essential not only for neurogenesis in early development but it ensures neuronal homeostasis throughout life. Moreover, deficits in many components of this pathway are linked to neurological disorders. Although the axonal retrograde transport of NT is a well-characterised process, molecular machinery controlling the sorting events is still poorly understood.
Recently, our lab discovered BICD1 motor adaptor protein as the key regulator of NT receptor sorting in embryonic stem cell-derived motor neurons (ES-MN). It led to hypothesis that this motor adaptor is a key factor regulating balance between receptor degradation and recycling which has an important implication on the downstream signalling pathways such as AKT and ERK1/2.
These findings, together with a still unpublished proteomic analysis of BiCD1-interacting proteins, led to development of this PhD project, which main goal is to further characterise the mechanism behind NT signalling and sorting. Elucidating the role of BICD1 and its novel binding partner – protein tyrosine phosphatase, non-receptor type 23 (PTPN23) in this process is the main aim of my PhD project.
Alex Fellows: Deficits in axonal transport as a target for pharmacological intervention in amyotrophic lateral sclerosis
The precise cause of motor neuron death in amyotrophic lateral sclerosis (ALS) is poorly understood. Previous work has shown that there are deficits in the axonal transport of signalling endosomes in embryonic motor neurons isolated from the SOD1G93A murine model of ALS compared to wild type. Correcting this deficit could therefore prove to be a valid therapeutic strategy. Insulin-like growth factor one receptor (IGF1R) has recently been identified as a mediator of retrograde axonal transport. This receptor tyrosine kinase plays an integral role in cell growth, survival and anti-apoptotic processes. The aim of my project is to explore the role of this receptor in axonal transport and during ALS disease progression. In order to do this, I will utilize both in vitro and in vivo models of ALS, including mouse models and neurons derived from patient induced pluripotent stem cells (iPSCs).
Group Project: Investigating the physiological role of nidogens
Nidogens are a family of basement membrane proteins with binding affinities for other key members of the extracellular matrix such as laminin, collagen IV and perlecan. They are currently believed to have a role in maintaining the structural integrity of the basement membrane. However, previous work in our lab identified a critical role for nidogens in binding of tetanus neurotoxin (TeNT) to the neuromuscular junction and its uptake into motor neurons (Bercsenyi et al., 2014). To continue with this work, a number of lab members are collaborating to investigate the receptor complex components, its transport dynamics and ultimately, the physiological role of internalised nidogen.
Ione Meyer: Exploring the role of nidogens at the neuromuscular junction
There are two nidogen proteins: nidogen-1 and nidogen-2. During early postnatal development there appears to be a preferential enrichment of nidogen-2 and a glycosylated form of nidogen-1 at the neuromuscular junction (NMJ). Preliminary results in our lab also suggest that the expression of nidogen-2 may not be uniform across all NMJs. The reason for this specificity is currently unclear. My project investigates the expression of nidogens and their associated proteins at the NMJs in different muscles. I will also aim to compare between muscles over development and in disease states. If differences at the NMJ are associated with particular muscle fibre types or motor neuron subtypes, this could have important implications for neurodegenerative diseases in which certain subtypes display specific vulnerability, such as amyotrophic lateral sclerosis (ALS).
Sergey Novoselov: Studying the tetanus toxin receptor complex by BioID approach
The BioID method exploits the property of the biotin ligase BirA to biotinylate proteins within a 10-nm radius. Fusion of BirA to an atoxic fragment of the TeNT (HcT) enables us to tag proteins coming into close contact with HcT during its recognition, internalisation and retrograde axonal transport in neurons. Tagged proteins are then isolated and identified by mass spectrometry and followed up by cell and molecular biology experiments to establish individual roles of these proteins in tetanus toxin propagation. Knowledge obtained by my research would be beneficial for combating tetanus as well as potentially useful for targeted neuronal drug delivery.
Sunaina Surana: Investigating the relationship between nidogens and neurotrophins
The tetanus neurotoxin is known to enter the same intracellular trafficking pathway entered by neurotrophic growth factors (Salinas et al., 2010). The recent finding that the tetanus toxin utilizes nidogens to facilitate its entry into neurons (Bercsenyi et al., 2014) raises the possibility that a similar mechanism might be in place to modulate the entry and sorting of neurotrophins. My project aims to study the role, if any, of nidogens on the uptake and transport of these growth factors. Proteins that are interacting partners of nidogens at the synaptic basement membrane are first identified using mass spectrometry. From this pool, neurotrophins are selected and, using a variety of biochemical and cell biological approaches, the effects of changing their binding status to nidogens are studied. Such an interaction with the extracellular matrix, if found, would be a stepping-stone for exploring the regulation of neurotrophins prior to their entry into neurons.
Andrew Tosolini: Characterising the mechanisms governing transcytosis of signalling endosomes in health and disease
Axonal transport ensures long-range delivery of essential components and signals between proximal and distal aspects of neurons. Transcytosis, the process where ligands are endocytosed in axon terminals and transported to the somatodendritic plasma membrane for release into the synaptic cleft, is another vital biological function that maintains neuronal survival. There is little knowledge of this biological process, however, this pathway is exploited by tetanus neurotoxin (TeNT). TeNT is retrogradely transported to motor neuron somata and is subsequently released into the synaptic cleft whereby TeNT binds to innervating interneurons, blocks their transmission and results in a spastic paralysis. My project will investigate the mechanisms governing transcytosis and in particular, the function of nidogens in the mechanisms responsible for sorting and transcytosis of signalling endosome cargoes. In addition, I will explore the role of transcytosis of signalling endosomes cargoes and pathological proteins in the spread of motor neuron disease/amyotrophic lateral sclerosis.
Deniz Tiknaz: Maintenance of the lab and embryonic stem cell facility
I have just joined the group in the beginning of March 2016. I am fortunate to be part of the team investigating axonal transport in motor neurons. My goal is to provide outstanding technical support for the project alongside my own professional development. My aspiration is to have a chance to realise my own ideas in the project once I picked up enough scientific background to intervene in this fascinating field of science.
Alex Rossor: Control of receptor trafficking as a therapeutic target in the hereditary neuropathies
My research interest is the inherited neuropathies. The inherited neuropathies are common diseases with a population prevalence of 1 in 2,500. They include a spectrum of genetic diseases including Charcot-Marie-tooth disease (CMT), hereditary motor neuropathy (HMN) and the hereditary sensory neuropathies. More than 70 disease causing genes have been identified, many of which are individually rare but collectively common. The inherited neuropathies frequently affect mobility and independence and in more severe cases can affect respiratory function leading to premature death. Despite their high prevalence and public health burden, there are no current treatments for the inherited neuropathies and this is contributed to by the large number of different disease genes.
A central theme of my research is that changes in endosomal sorting at any level can lead to changes in endosome to plasma membrane recycling and alter the cell surface receptor landscape of neurons. Changes in the cell surface receptor landscape can in turn lead to activation of intracellular pathways that are detrimental to the cell and that the identification of these cell surface receptors may provide a readily accessible therapeutic strategy. Whilst this fellowship will focus on one type of hereditary neuropathy, almost all inherited neuropathy disease genes can be predicted to affect at least one aspect of endosomal sorting and may also result in altered cell surface receptor expression. This central theme has long been used in the field of cancer where research has focused on changes in cancer cell surface receptor expression. Such an approach has transformed cancer therapeutics.
James Sleigh: Dissecting Neuronal Specificity in Hereditary Neuropathy
I am particularly interested in understanding the cause of neuronal susceptibility often observed in diseases that affect the peripheral nervous system. Motor and sensory nerves appear to be highly susceptible cells; mutations in numerous different genes important throughout the body manifest in a very specific detrimental effect on these peripheral neurons.
Charcot-Marie-Tooth disease (CMT) is a large group of genetically diverse peripheral neuropathies that share the principal pathological feature of progressive motor and sensory impairment. CMT type 2D (CMT2D) is caused by dominant, toxic gain-of-function mutations in a gene called GARS, which encodes glycyl-tRNA synthetase (GlyRS). GlyRS attaches the amino acid glycine to its cognate transfer RNA (tRNA), thereby priming the tRNA for protein translation. This housekeeping function of glycine aminoacylation explains the widespread and constitutive nature of GARS expression, but how do mutations that affect a protein found in all cells selectively trigger peripheral nerve degeneration?
I currently have a Wellcome Trust Sir Henry Wellcome Postdoctoral Fellowship that I am using to answer this question. I am using live imaging of cellular dynamics, including axonal transport, to interrogate both the motor and sensory systems in cell and animal models to better understand the cause of neuronal fragility seen in CMT2D.
Alison Twelvetrees: The coordination of microtubule motors during axonal transport
Microtubule motor proteins, such as cytoplasmic dynein and kinesin-1, are responsible for long range transport of cellular cargoes. The length and complexity of neurite networks makes neurons uniquely reliant on this type of long-range intracellular transport and many neurological diseases display defects in this transport; including Huntington's Disease, Alzheimer's Disease and Amyotrophic Lateral Sclerosis. However, many fundamental aspects of microtubule transport and the regulation of motor proteins remain unknown. My long-term research interests focus on understanding how different microtubule motor proteins coordinate with each other to ensure healthy neuronal development and function, and how defects in these processes contribute to neurological disease.
My current work has outlined a molecular mechanism for the transport of cytoplasmic dynein from soma to axon terminal, driven by direct interactions with the anterograde motor kinesin-1. This work used biochemistry, single-molecule in vitro assays and live cell imaging assays to investigate the underlying mechanism. This lead us to propose a model for the axonal transport of cytosolic cargos, based on short-lived direct interactions of cargo with a highly processive anterograde motor. I am currently using CRISPR/Cas9 gene editing technology to develop embryonic stem (ES) cell lines that express knock-in reporters, so that movements of motors can then be analysed in neurons differentiated from mutant ES cells.