Mutations in Munc18-1 are associated with three infantile epileptic encephalopathies, but the mechanistic relationship between mutations and these diseases is unknown. The long term goal is to clarify mechanisms by which specific synaptic dysfunctions trigger neurological disorders. The objective in this application is to determine how mutations in Munc18-1 cause neuronal defects, and to identify rescue strategies to reverse such deficits. Munc18-1 (also called STXBP1, SEC1 in yeast, Rop in flies, and unc18 in worms) controls neurotransmitter release at the synapse via binding to multiple effector proteins. Over 30 heterozygous de novo mu- tations have been identified in the Munc18-1 gene that cause the infantile epileptic encephalopathies Ohtahara, West, and Dravet syndrome, but it is unknown how these mutations affect neurons and trigger three different diseases. The central hypothesis, based on strong preliminary data, is that mutations in Munc18-1 result in defects in its folding, stability and localization, and elicit same defects in wild-type Munc18-1. This loss of functional Munc18-1 subsequently impairs the function of its effectors and triggers synaptic dysfunction, which can be restored by stabilizing Munc18-1. The rationale for these studies is that understanding of how mutations in Munc18-1 trigger synaptic dysfunction in infantile epileptic encephalopathies will create opportunities for the development of novel therapies beyond the current, limited symptom-based therapy. Guided by strong preliminary data, this hypothesis will be tested in three specific aims: 1) Determine the impact of disease-relevant mutations in Munc18-1 on protein stability; 2) Determine how Munc18-1 mutants affect the stability of its effectors and synapse function; and 3) Identify rescue strategies to stabilize Munc18-1 and restore its function. Under the first aim, stability, folding, aggregation and intracellular targeting will be quantified for Munc18-1 wild- type and mutants, combining purified recombinant proteins and primary neurons with biochemical and cell biological techniques. Under the second aim, stability, interaction, and targeting of Munc18-1's effectors syntaxin- 1, Doc2, Mint1, Mint2, and rab3, as well as synapse integrity and function will be analyzed, using purified proteins, primary neurons, and in vivo mouse and worm models. Under the third aim, chemical and molecular chaperones will be employed to restore deficits in Munc18-1 and in synapse structure and function, using same paradigms as for aims 1 and 2. This research is significant, because it will clarify the molecular mechanisms underlying Munc18-1-linked epilepsies, and will have translational importance in the development of new rational treatments. This research is innovative, because it 1) tests the novel hypothesis that Munc18-1 mutations cause synaptic dysfunction via a dominant-negative mechanism, 2) uses a multidisciplinary and systematic approach that has not previously been used in this research area, 3) is technically innovative because of newly generated C. elegans strains, and 4) shifts focus from a symptom-centered perspective to an approach that focusses on understanding convergent underlying disease mechanisms that pivot on Munc18-1.

Public Health Relevance

The proposed research is relevant to public health because three infantile epileptic encephalopathies are related to mutations in the Munc18-1 gene, and understanding how these defects cause disease will allow us to develop a novel intervention therapy. Thus, this research is relevant to the NIH's mission that pertains to supporting the ongoing basic research, foster research that has immediate medical relevance, and encourage translational research that links the discoveries from basic research into therapeutic interventions for treating neurological disorders such as epilepsy.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Therapeutic Approaches to Genetic Diseases Study Section (TAG)
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Whittemore, Vicky R
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Weill Medical College of Cornell University
Schools of Medicine
New York
United States
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Guiberson, Noah Guy Lewis; Pineda, André; Abramov, Debra et al. (2018) Mechanism-based rescue of Munc18-1 dysfunction in varied encephalopathies by chemical chaperones. Nat Commun 9:3986