NIHRDataInsights/HRCSData huggingface dataset:
Title: Roles of intron retention and splicing factors in axons
Funder: Wellcome Trust

Abstract: In recent years, RNA dynamics in axons and dendrites have arisen as central to neuronal maturation and maintenance of healthy neuronal connectivity. However, the functional roles of transported mRNAs and their binding proteins are scarcely understood. Some of the proteins associated to local mRNAs are splicing factors and spliceosome proteins. We recently demonstrated that their axonal role is complex, as, in developing neurons in vivo, the splicing factor SFPQ and the U1 spliceosome protein snRNP70 locally control axonal shape and connectivity, through modulation of the local transcriptome. These findings open two essential questions: how are these splicing proteins shaping the transcriptome landscape locally and what is the molecular nature of the axonal interactions between specific mRNAs and these two proteins? Moreover, the recent findings of axonal and dendritic partially spliced mRNAs, retaining a single intron, open the additional question of the role(s) of intron-retaining transcripts in neurites and the possibility of direct functional interaction between retained introns and splicing proteins. We propose to answer these essential questions, at cellular and molecular levels, using zebrafish developing neurons (in embryos and in culture) as experimental models.

Lay abstract: In our nervous system, neurons transport mRNAs (transcripts) along axons and dendrites, where they are translated into proteins, centimeters away from their cell body. Dysfunctions in this mechanism are associated to human neurological and neurodegenerative disorders. We recently discovered that the diversity of axonal transcripts depends upon two axonally-located splicing proteins. Yet, these proteins were so far known for transforming draft mRNA into mature transcripts in the cell nucleus, their role on mRNA in neurites being unexplored. We also found that some axonal mRNAs are partially immature (still containing a piece of "draft" RNA called intron). We propose to i) understand how splicing proteins modulate axonal mRNA diversity, ii) reveal what introns do in axons, and iii) discover the functional links between introns and splicing proteins in axons. We will answer these essential questions, at cellular and molecular levels, using zebrafish animals and primary neuronal cultures as experimental models.

Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: GENERIC HEALTH RELEVANCE;NEUROLOGICAL
Title: Deciphering morphogenetic cues encoded in cell shape
Funder: Wellcome Trust

Abstract: Tissue morphogenesis is associated with dramatic changes in cell morphology. Whilst these cell shape changes may define final tissue form, how they impact and/or direct other key morphogenetic processes remains unclear. I aim to redefine our understanding of the role of cell shape in tissue formation by revealing that, far from simply a structural feature, cell shape encodes critical instructive information that directs diverse morphogenetic events. Using the vasculature and neural crest as morphogenetic models and combining zebrafish live-cell imaging, in-vitro micropatterning and computational modelling, this proposal will investigate (1) if acquisition of specific cell geometries in interphase determines whether cells will divide symmetrically or asymmetrically to direct post-mitotic cell fate, (2) whether signal-induced cell shape changes create positive-feedback that amplifies signal to expedite fate decisions, and (3) how local cell shape remodelling, driven by polarised mRNA targeting and translation, functions to orient motile cell polarity and tissue movements. As such, this work will determine if cell shape unexpectedly encodes diverse morphogenetic cues that direct fundamental decision-making processes underpinning tissue building. Considering that cell shape change is an inherent feature of almost all forming tissues, mechanisms uncovered here may be broadly conserved and of therapeutic relevance in numerous tissue contexts.

Lay abstract: During tissue development, growth and repair, cells undergo extensive morphological changes. Whilst these cell shape dynamics clearly sculpt tissues, how they impact other key cellular events that drive tissue morphogenesis is unclear. My work suggests that cell shape unexpectedly directs multiple cellular processes, such as cell division, signalling and polarity, to function as a synergistic modulator of critical cell fate and behaviour decisions underpinning tissue building. Using a multidisciplinary approach integrating in-vivo live imaging, gene-editing and computational modelling techniques, I aim to mechanistically understand how cell shape achieves this by defining; (1) how interphase cell morphology directs asymmetric versus symmetric cell division/fate, (2) how signal-induced cell shape changes create positive-feedback that amplifies signalling responses and (3) how local cell shape remodelling polarises motile cells/tissues. Hence, I will define cell shape as a novel means to encode and process diverse instructive cues that direct key decision-making processes during tissue formation.

Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: GENERIC HEALTH RELEVANCE;NEUROLOGICAL
Title: Determining the role of activity-dependent bulk endocytosis via new molecules
Funder: Wellcome Trust

Abstract: Synaptic vesicle (SV) endocytosis is essential for the maintenance of neurotransmission, particularly during intense neuronal activity. Under these conditions the dominant endocytosis mode is activity-dependent bulk endocytosis (ADBE), suggesting it should perform a central role. However determining the physiological role of ADBE has been hindered by a limited understanding of its molecular mechanism. We propose to address these challenges by discovering presynaptic molecules with a specific and selective role in ADBE. This will be achieved by integrating functional proteomics, a spectrum of complementary SV recycling assays and detailed molecular studies to establish a mechanistic basis for key steps in ADBE. This will facilitate generation of transgenic rodents and molecular tools to disrupt / enhance specific stages of ADBE in vivo. Importantly, this research strategy is already established with a series of presynaptic molecules discovered that have specific roles in ADBE. We will exploit these findings to determine how ADBE impacts on neurotransmission at the synaptic, neuronal, circuit and behavioural level. This work will reveal fundamental mechanisms that underpin neurotransmitter release during high intensity firing, and potentially ADBE-specific interventions that will manipulate presynaptic performance exclusively during intense neuronal activity.

Lay abstract: Brain cells (neurones) communicate by releasing chemical neurotransmitters from small compartments called synaptic vesicles (SVs). The maintenance of neurotransmitter release is dependent on SVs being reformed by a process called endocytosis. There are different mechanisms of endocytosis, and during very high brain activity the dominant mechanism is activity-dependent bulk endocytosis (ADBE). A number of essential processes are triggered by high brain activity, such as generation of learning and memory, suggesting ADBE will be an important mechanism in these events. However research progress in this direction has been hindered by the absence of identified molecules that are specific to ADBE. In this application we propose to discover new ADBE-specific players, determine how they work and then disrupt their normal function to discover how ADBE controls neurotransmitter release, brain communication and animal behaviour. Importantly, this process is already underway and we have a series of candidate ADBE molecules to investigate. This work will reveal how neurones communicate during high brain activity and critically how this communication could be altered by manipulating ADBE. This work therefore has the potential to allow a specific and selective intervention to modulate brain communication in healthy individuals or restore normal function in neurological disease.

Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: GENERIC HEALTH RELEVANCE;NEUROLOGICAL
Title: Protein-protein interactions in the early stages of amyloid assembly mechanisms
Funder: Wellcome Trust

Abstract: How and why proteins aggregate is an important fundamental question. It also has far-reaching biomedical importance given the increasing prevalence of amyloid-diseases in today’s ageing population. Whilst some amyloid precursors are intrinsically disordered, others are natively folded. Each must undergo major conformational changes to form amyloid fibrils. Defining these changes is key to developing therapeutic strategies. This proposal aims to achieve this by focusing on two overarching questions: 1. What is the nature of the protein-protein interactions that initiate amyloid formation; 2. Can we use this knowledge to develop molecules able to control aggregation in vitro and in vivo? The challenges in answering these questions lie in the heterogeneity of aggregating species, their transient/weak interactions, and the fact that amyloid precursors adopt non-native, dynamic structures. We will meet these challenges: exploiting cutting edge structural methods to map the protein-protein interactions that commit proteins to aggregate and will use the knowledge gained to inform the design of small molecules/artificial proteins able to control aggregation by targeting these surfaces. Focusing on islet-amyloid-polypeptide (involved in type II diabetes) and &beta;2-microglobulin (involved in systemic amyloidosis) the goal is to enhance our fundamental understanding of protein aggregation and to inspire new strategies for intervention in disease.

Lay abstract: Over 50 human disorders involve the aggregation of proteins into amyloid plaques. These disorders include type II diabetes, Alzheimer’s and Parkinson’s diseases, as well as a host of rarer disorders, for all of which there is currently no cure. The need to develop successful therapies for amyloid disease is a major priority for human health today. Amyloid formation is thus of intense interest both to academics focussed on how changes in the sequence of a protein causes amyloid formation, and to pharmaceutical companies who require this knowledge to design drugs able to prevent disease. Here I propose to focus on two amyloid diseases - type II diabetes and dialysis-related amyloidosis - which involve the formation of amyloid deposits in different sites in the body. By combining the powers of structural molecular biology and chemical biology we will map the early processes that initiate amyloid formation and use the knowledge gained to create molecules able to control assembly by targeting these sites. By testing the effects of this toolkit of reagents in cells and in model organisms our vision is to reveal new understandings of why proteins aggregate and to develop new strategies to control aggregation-induced cellular dysfunction and disease.

Research activity codes: 5.1 PHARMACEUTICALS
Health categories: GENERIC HEALTH RELEVANCE;NEUROLOGICAL
Title: Protein quality control in health and disease
Funder: Wellcome Trust

Abstract: Accumulation of misfolded proteins is a defining feature of numerous diseases. My work aims to identify strategies that rescue cells from protein quality control failure with the view that such strategies may lead to generic therapies for diverse protein misfolding diseases. Recent and transformative discoveries in the lab have validated this concept. One recently discovered strategy consists of a selective inhibitor of the constitutive regulatory subunit of the eIF2&alpha; phosphatase to transiently decrease protein synthesis rates, thereby correcting folding defects by increasing protein quality control capacity. This approach is safe, novel and very promising because it may benefit common neurodegenerative diseases. This proposal aims at testing this new and exciting possibility in mouse models of common neurodegenerative diseases, reconstituting phosphatase inhibition using purified components and examining the functional consequences of the inhibitors on the translatome. We will also identify other phosphatases controlling the termination of stress signalling pathways anticipating that prolonging stress signalling could be a generic paradigm to enhance the natural cellular defence systems. This research programme will bring fundamental discoveries with a translational potential relevant to a group of devastating and so far incurable diseases.

Lay abstract: Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, Amyotrophic Lateral Sclerosis and Prion disease are devastating and affect an increasing number of individuals in ageing populations. Each disease is caused by the progressive dysfunction and death of specific nerve cells in selective regions of the brain due to the accumulation of proteins of aberrant shape. My group has identified selective, safe and potent small molecule inhibitors that enhance the natural defence systems against misshapen proteins. In mice, the inhibitor protects against Huntington’s disease. In this work, we will examine the benefit of our inhibitors in models of common neurodegenerative diseases, characterize their mode of action in vitro and in cells and search for other targets that could be inhibited to enhance different cellular defence mechanisms. Our work is innovative because it targets phosphatases, a class of enzymes that were thought to be undruggable. We will manipulate fundamental cellular processes to correct the cellular dysfunction at the origin of diverse diseases. Ultimately, the knowledge and the inhibitors emanating from our work may be used to develop treatments for a group of devastating diseases.

Research activity codes: 5.1 PHARMACEUTICALS
Health categories: GENERIC HEALTH RELEVANCE;NEUROLOGICAL
Title: Towards a thorough mechanistic understanding of endosomal cargo sorting.
Funder: Wellcome Trust

Abstract: An intracellular nexus for regulating the membrane trafficking of many of the 5,000+ integral proteins encoded by the human genome is the endosomal network. Composed of vesicular and tubular early and late endosomes, the network’s principal role is to sort integral proteins (termed ‘cargoes’) arriving from the cell surface and the biosynthetic pathway between two fates: either sorting to the lysosome for degradation, or retrieval from this fate for export to the cell surface, the biosynthetic pathway or other specialised organelles. Whilst the molecular details of degradative cargo sorting have been well documented, those events that conduct cargo retrieval and export remain poorly understood. Our research has sought to fill this fundamental void in metazoan cell biology. A master conductor of endosomal retrieval and export is the retromer pathway - in human cells this orchestrates the sorting of >900 cargoes. Establishing how this pathway functions is central to understanding the evolution, organisation and activity of endosomal sorting. With Wellcome support we will address two integrated questions: • The fundamental question of how the retromer pathway is organised and integrated with other pathways to orchestrate global endosomal cargo sorting. • How understanding of retromer pathway function may provide vital insight into neurodegenerative diseases.

Lay abstract: For over ten years my laboratory has been identifying the mechanistic basis by which proteins and lipids are sorted through a specific aspect of the human cell's intracellular membraneous maze, the endosomal network. We have studied a number of ancient, highly conserved protein complexes that act as molecular machines to orchestrate protein and lipid transport through this network. One of these is the retromer complex. Our research, and that of others, is defining retromer function and is revealing its importance in a variety of cellular processes that are vital for normal cell function. Furthermore, it has become apparent that defects in retromer underlie a variety of human diseases including age-related neurodegenerative diseases such as Parkinson disease and Alzheimer disease. With Wellcome funding we will address new fundamental questions arising from this research, thereby providing greater fundamental insight into retromer's many roles in normal physiology and its deregulation in pathophysiology.

Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: GENERIC HEALTH RELEVANCE;NEUROLOGICAL
Title: Regulation and function of PINK1 kinase in Parkinson's disease
Funder: Wellcome Trust

Abstract: Loss-of-function mutations in PTEN-induced kinase 1 (PINK1) cause familial Parkinson's disease. My laboratory has contributed mechanistic understanding into how PINK1 functions in a common signalling pathway with the ubiquitin E3 ligase Parkin to regulate mitochondrial quality control. Highlights of our past work include the unexpected discovery that PINK1 can phosphorylate ubiquitin and the recent crystal structure of insect PINK1. To date it remains unknown whether our current understanding of PINK1 biology is conserved in neuronal cell types and tissues of physiological relevance to Parkinson's. To address this, we will employ quantitative proteomics to uncover novel substrates of Parkin in primary mouse neuronal systems as well as tissues of a Parkin Ser65Ala knock-in mouse model, we have characterised, that exhibits mitochondrial dysfunction in vivo. We will investigate how mutations in other Parkinson's genes particularly VPS35 regulate PINK1 signalling. We will investigate the mechanism by which human PINK1 becomes activated including solving the crystal structure of full length human PINK1 bound to its substrate ubiquitin. These analyses will provide a framework of knowledge that will be critical for the development and evaluation of PINK1 activators that are currently being explored as a potential therapy against Parkinson's.

Lay abstract: Parkinson's disease is an incurable disorder of the brain in which brain cells are progressively lost. Mutations in two genes PINK1 and Parkin cause heritable forms of Parkinson's. Our laboratory has made major advances in understanding how PINK1 and Parkin communicate closely to maintain the health of the 'batteries' of cells called mitochondria. However, we still have poor understanding of the details of this communication in the brain cells relevant to Parkinson's. We plan to study brain cells from mice and and apply state-of-the-art methods in mass spectrometry and biochemistry to identify new targets of PINK1 and Parkin. We also plan to see how other Parkinson's genes affect PINK1 and Parkin. Finally, we will investigate how PINK1 is controlled by cellular stress. The new knowledge elaborated in this study will be important for the development and testing of drugs targeting PINK1 and Parkin as potential therapies against Parkinson's.

Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: GENERIC HEALTH RELEVANCE;NEUROLOGICAL
Title: RNA metabolism in developing neurons
Funder: Wellcome Trust

Abstract: Understanding how neurons translate extracellular cues into specific patterns of gene expression is amongst the major goals of modern neurobiology. Efforts to unravel the molecular basis of localised gene expression have mostly focused on transcriptional control and mRNA transport. However, post-transcriptional mRNA processing is increasingly recognised as an essential step in the propagation of genetic information. Importantly, incorrect processing and delivery of mRNA causes developmental defects and severe human neurological disorders. The overarching scope of this project is to investigate novel mechanisms of mRNA metabolism that are likely to be conserved in all mammalian cells and to provide new insights regarding how gene expression is regulated in neurons. The 3’ untranslated regions (3’UTRs) of RNA transcripts play a key role in mediating many steps of RNA metabolism, including splicing, nuclear export and localization. We performed 3’end-Seq screens of transcripts localised in sub-cellular compartments of sympathetic neurons and are uniquely positioned to tackle the question of how 3’UTRs regulate RNA metabolism in neurons. We will address the following questions: 1. What determines the fate of a bifunctional mRNA? 2. Does the 3’UTR influence the proteome in sympathetic neurons? 3. Do 3’UTRs generate a new class of RNAs that regulate neuronal development?

Lay abstract: Neurons are highly polarised cells that communicate electrical signals between each other and between our tissues and the brain. Thus, they often need to be of considerable length and require specialised transport mechanisms to communicate between the far ends of the cell. It is now clear that transport, metabolism and local translation of mRNAs are critical for the growth and health of neurons such that it has been hypothesized that many neuronal disorders are due to alterations of these processes. In a systematic approach that spans from molecular analyses to the investigation of nerve growth in vivo, we will identify the biochemical mechanisms that regulate mRNA metabolism in neurons and how they influence neuronal growth and development. The highly innovative nature of our research will provide new and exciting findings that will have implications for the health of the nervous system and for understanding the basis of neuronal diseases.

Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: GENERIC HEALTH RELEVANCE;NEUROLOGICAL