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BDSRA 2015 CLN1 Wishart


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2015 CLN1 Wishart

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BDSRA 2015 CLN1 Wishart

  1. 1. Wishart Laboratory : Infantile Batten Disease – A synaptic Study Maica Llavero Hurtado1, Tom Gillingwater1, Giusy Pennetta1, Jon Cooper2 & Tom Wishart1 1 University of Edinburgh, 2 Kings College London, United Kingdom. Contact: (Neurobiology Division, The Roslin Institute, University of Edinburgh) Informational Text Here Acknowledgements: Dundee Proteomics Facility, Members of the Pennetta, Cooper and Wishart labs. Maica Llavero is funded by the Darwin Trust, Tom Wishart is funded by the BBSRC and MRC. 1. What is the problem? DNA  muta)on   Non-­‐func)onal   CLN1  protein   Neuron  degenera)on   2. Why are synapses important? …a  cell  body   …an  axon   …and   synapses   A  neuron  has…   5. Identification of protein differences in synapses B  Thalamus  &  Cortex   dissec:on   Isola:on  of   synapses     A   1 month 3 month 5 month 7 month Late-symptomaticEarly-symptomatic Disease  onset?   controls   controls   controls   controls  cln1/-­‐   cln1-­‐/-­‐   cln1-­‐/-­‐   cln1-­‐/-­‐   Protein   extrac:on       C   Protein  iden:fica:on  and   quan:fica:on       D   Protein  A   Protein  B   6. Identification of disease regulators in other animal models Healthy  eye   Healthy  Fruit  Fly   BaGen  Disease  Fruit  Fly   Small  and   degenera:ve  eye   Healthy  synapse   Disrupted  synapse   Protein  A   Protein  B   We  can  modify  the   levels  of  our  protein   candidates  (iden:fied   in  5)  in  a  fly  model  of   BaGen  Disease  to  see  if   we  can  change  disease   progression.   Modifying   Protein  A  levels   we  make  the   eye  bigger.   Makes  disease   beGer.   Modifying   Protein  B  levels   we  make  the   eye  smaller.   Makes  disease   worse.   7. What does this means for therapy? Biochemistry   techniques   Small  animal   models   Large  animal   models   Human   Can  we  target   our  candidates   with  drugs?   Can  these  drugs   become  a  therapy  for   neurodegenera:on?   The brain is a very complex organ. It contains billions of cells called neurons. Neurons form a very tight network of connections. When this network is disrupted it can cause a wide range of different diseases. In Batten disease, synapses (communication points between nerve cells/neurons) begin to break down early in disease progression. The reasons why synapses are so vulnerable is unknown. Our laboratory is trying to work out what mechanisms govern the vulnerability of synapses and could therefore be important in regulating disease progression. Neurons appear to be quite complicated cells. There are many types of neurons but they all have synapses. Synapses are e s s e n t i a l c o n n e c t i o n s e n a b l i n g communication between neurons. Their stability is essential for normal brain function. Synapses break down at very early disease stages in many neurodegenerative conditions including Alzheimer’s, Huntington’s, Motor Neuron and Batten diseases. It is therefore imperative to understand the role of synapses in disease. 4. What happens in disease? 1 month 12 month 3 years 10 years Late-symptomaticEarly-symptomatic Disease  onset?   •  re:nal  degenera:on   •  blindness   •  cogni:ve  and  motor   deficits   •  seizures   •  flat  EEG     •  Premature   mortality       1 month 3 month 5 month 7 month Late-symptomaticEarly-symptomatic Disease  onset?   synap:c   breakdown   beginning  in   thalamus   •  Seizures   •  Death   Mouse  Model   Human   synap:c   breakdown   beginning  in   cortex   Mouse  brain   The brain can be subdivided into different areas by the cell types contained within and functions performed by that portion of the brain. Not all brain regions are affected at the same time in neurodegenerative disease. For example, in Alzheimer’s hippocampus goes first. In Huntington’s it is striatum. This is also true for Batten Disease. In the mouse models available for the disease, synaptic breakdown is first detected in thalamus and then later in cortex. This raises the question: what makes some synapses more vulnerable than others? CLN1   3. What is CLN1 doing in the synapse? CLN1 protein is not specific to neurons. It is involved in the function of lysosomes which are part of the machinery for waste clearance present in every cell. If lysosomes and therefore CLN1 are present in all cell types why does its loss have such a profound effect on neurons, presenting as a neurodegenerative disease? The presence of CLN1 protein must therefore have different consequences in different tissues and be particularly important for the maintenance of neurons. We therefore need to know what CLN1 interacts with in neurons and their synapses. We use what is known about synaptic breakdown in CLN1 mouse models to examine how the composition of synapses changes due to the loss of CLN1 and throughout disease progression. We need protein extractions from mouse brain, specialised equipment, powerful computers and complex software to create and analyze the data. This workflow allows us to infer what CLN1 interacts with in synapses (see 3 above), to identify proteins which could control the stability of synapses (see 4 above) and regulate disease progression (see 6 next). E   Correla:on   We can model Batten disease in flies. Flies also have synapses which break down with Batten like mutations. However, their most obvious effect are alterations in the eye. Whilst they are obviously not as complex as humans, they are useful for testing candidates for their potential in therapy. After identifying interesting protein candidates we target them in our Batten disease flies. By modifying the levels of our protein candidates (identified in 5) we see if they can make the Batten disease eye fly better. We can successfully identify proteins changed in synapses due to loss of CLN1 which can change the disease in Batten flies. These results are extremely preliminary and testing of candidates is needed in more “biologically relevant” larger models before we can be sure they well be useful for humans. However, these findings are an important proof of principle because it means that by starting with what is happening in synapses we can now search for candidates which change the rate of degeneration in Batten. It also means that successful candidates may also be effective in a range of neurodegenerative diseases where synapse are early pathological targets. We are here Some of the generic schematics presented here are modified from the internet.