Identifying endosomal/lysosomal components in Drosophila: modeling lysososomal storage disease

Lead Research Organisation: University of York
Department Name: Biology

Abstract

Lysosomal Storage Diseases (LSDs) are a set of genetic diseases in humans that cause a major disruption of the late endosome (LE), a compartment of the cell where components of the cell are sent to be degraded. This LE disruption leads commonly to a severe and debilitating neurodegeneration and premature death during childhood. I am proposing to use the advanced genetics of the model organism Drosophila melanogaster as a system for identifying novel LSD genes and the molecular signaling pathways that are perturbed in LSDs. I intend to do this to further our understanding of the LE and LSDs since many of the genes encoded by LSD loci are important to LE function. D. melanogaster can manifest all of the LSD symptoms. This fact, allied to the advanced genetics available in this organism plus publication of its genome sequence makes work rapid, novel and informative to the understanding and treatment of LSDs in humans.

Technical Summary

Modern molecular genetics has allowed the cloning of many of the genes underlying lysosomal storage diseases (LSD) and this set of diseases have in turn revealed many insights into the normal cell biology of the late endosomal/lysosomal (LE) system. The LSDs a a set of untreatable recessive diseases in humans that result in LE expansion, lipofuscin deposition, neurodegeneration and early death. In the following proposed work I intend to identify novel LSD loci in the genetically tractable organism Drosophila melanogaster to futher our understanding of LSD at a molecular and pathological level. This will be achieved by using a novel, rapid and high through-put genome wide RNAi screen in tissue culture cells in conjunction with a simple assay for LE dysfunction based on a known LSD phenotype. Loss of gene function giving rise to an LSD phenotype is likely to identify resident LE proteins or proteins involved LE function that are important to LSD pathology. The second section of proposed work develops previous findings where I (and others) identified the mutant spinster as an LSD loci in Drosophila (Nakano et al., 2001; Sweeney and Davis, 2002). Mutants of spinster have overgrown larval neuromuscular synapses, an expanded LE, lipofuscin deposition and shortened life-span. Using the neuromuscular synaptic overgrowth of spinster mutants as a read-out of LE dysfunction I propose to manipulate known components of the LE and identify the relationship to known components of the LE and the cellular signaling defects caused by this LSD. In a third aim, I intend to use a novel genetic screen to identify the signaling pathways downstream of LE dysfunction that leads to the cellular pathology of LSDs. These three approaches will identify molecules and mechanisms that are key to the control and organization of the LE as it relates to LSD pathology.

Publications

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Ahmad ST (2009) Genetic screen identifies serpin5 as a regulator of the toll pathway and CHMP2B toxicity associated with frontotemporal dementia. in Proceedings of the National Academy of Sciences of the United States of America

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Milton VJ (2012) Oxidative stress in synapse development and function. in Developmental neurobiology

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Milton VJ (2011) Oxidative stress induces overgrowth of the Drosophila neuromuscular junction. in Proceedings of the National Academy of Sciences of the United States of America

 
Description Alzheimers Society PhD studentship
Amount £27,000 (GBP)
Organisation Alzheimer's Society 
Sector Charity/Non Profit
Country United Kingdom
Start 03/2010 
End 03/2013
 
Description Project Grant
Amount £192,000 (GBP)
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 09/2010 
End 08/2013
 
Title Drosophila models of Fronto-temporal Dementia, Lowe's syndrome, Hereditary Sensory Neuropathy type 1, saposin deficiency and Mucopolysaccharidosis type IIIB 
Description We have generated in Drosphila five novel neurological disease models associated with defective membrane function: A knockout of OCRL1, a model of Oculocerebrorenal syndrome of Lowe, a knockout model of saposin deficiency (a lysosomal storage disease model), a functional model of Hereditary sensory neuropathy type 1 and a model of Sanfilipo syndrome (mucopolysaccharidosis type IIB, a lysosomal storage disease). Lastly, we have developed a screenable model of Fronto-temporal dementia associated with chromosome 3. 
Type Of Material Model of mechanisms or symptoms - non-mammalian in vivo 
Year Produced 2006 
Provided To Others? Yes  
Impact We have initiated a collaboration with Dr Fen Biao-Gao of the Gladstone Institute, University of California, San Francisco investigating the model of FTD3. This resulted in a publication in PNAS describing the involvement of the innate immune response in frontotemporal dementia. We have also used our model of Sanfilipo Syndrome IIIB to initiate a collaboration with Dr Keith Stubbs of the University of Western Australia to develop chemical chaperones for protein folding therapies in this neurodegenerative disease. 
 
Description Analysis of BicD function at the synapse 
Organisation Medical Research Council (MRC)
Department MRC Laboratory of Molecular Biology (LMB)
Country United Kingdom 
Sector Academic/University 
PI Contribution Our lab provided the TEM analysis of stimulated and unstimulated synapses. We also conducted morphological analysis of BicD mutants. This gave a description of BicD as a novel regulator of clathrin dynamics at the synapse/
Collaborator Contribution Publication of Li et al (2010) EMBO J.
Impact We published a paper in EMBO J showing a novel role for the cargo adaptor protein BicD at the synapse. We will continue the collaboration to further dissect the role of this protein at the synapse.
Start Year 2007
 
Description Modeling Frontotemporal dementia in Drosophila 
Organisation University of California, San Francisco
Department Gladstone Institute of Neurological Disease
Country United States 
Sector Academic/University 
PI Contribution Performance of genetic screen and identification of enhancers of Frontotemporal dementia model
Collaborator Contribution Publication outcome, Ahmad et al.
Impact Successful application for an Alzheimer's Society PhD studenthsip for 3years and recruitment of student. Publication of findings in PNAS (Ahmad et al, 2009) of involvement of immune response in CHMP2B model of Frontotemporal dementia model in Drosophila
Start Year 2006
 
Description Oxidative stress and synaptic growth 
Organisation University of Cambridge
Department Department of Zoology
Country United Kingdom 
Sector Academic/University 
PI Contribution Data arising from the MRC grant and published in Milton et al., 2011 led to a grant application to the BBSRC jointly with Dr Matthias Landgraf. Our lab examine oxidative stress effects on neuromuscular junction synapses and Dr Landgraf's group examine central synapses.
Impact Award of BBSRC Regional Schools Champion (£1600 over two years) for outreach activity.
Start Year 2010
 
Description Appearance on Local Radio 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach Local
Primary Audience Media (as a channel to the public)
Results and Impact 10 minute appearance talking about the work on oxidative stress and synapse growth on local BBC Radio York breakfast show.

I have been asked back to contribute regularly to the breakfast show, to discuss newspaper headlines and scientific progress. As a result of the appearance, articles appeared on the BBC North Yorkshire website and the Yorkshire Post newspaper.
Year(s) Of Engagement Activity 2011
 
Description Dementia Lay Meeting 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Talk by PhD student to Northern Section of Alzheimer's Society Lay Members. Only scientific talk of the day.

Repeated requests for copies of the presentation, contact with lay members offering support and asking questions
Year(s) Of Engagement Activity 2012
 
Description Outreach Activity at University of York 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Schools
Results and Impact 30 pupils involved in a practical demonstration activity on nerve function and sensory system function

Currently writing a small teaching paper to be submitted to Invertebrate Neuroscience describing a simple, cheap and reliable practical to teach sixth-form students or undergraduate students about sensory function in Drosophila larvae. Practical is now available 'off the shelf' and will be delivered regularly to other sixth-form cohorts locally.
Year(s) Of Engagement Activity 2010
 
Description Oxidative Stress coverage in local press 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Press reports on the publication of our paper in PNAS, the York Press, the Yorkshire Post.

Contact from public
Year(s) Of Engagement Activity 2011
 
Description School visit - York 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? Yes
Geographic Reach Regional
Primary Audience Schools
Results and Impact Sharing of research skills with secondary pupils

Greater interest from Biology students in the Biology Degree at York.
Year(s) Of Engagement Activity 2008
 
Description School visit to York 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Type Of Presentation Workshop Facilitator
Geographic Reach Regional
Primary Audience Schools
Results and Impact Workshop on pain developed by Dr Sweeney (and published as a teaching paper) using Drosophila larvae. Attended by 30 sixth form students

School has been returning on a yearly basis, the activity supports core curriculum areas in neurobiology.
Year(s) Of Engagement Activity 2012
 
Description School visit2 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Type Of Presentation Keynote/Invited Speaker
Geographic Reach Local
Primary Audience Schools
Results and Impact Talk to local girl's school about our work on Frontotemporal Dementia. Followed this with talk on Science as a career

Contact now made with school for work shadowing and outreach activities in teaching labs at York
Year(s) Of Engagement Activity 2012