Project

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JUMP: Junction-specific mapping of interorganellar sensors and their signaling activities

English title JUMP: Junction-specific mapping of interorganellar sensors and their signaling activities
Applicant Aye Yimon
Number 190192
Funding scheme Spark
Research institution Institut des sciences et ingénierie chimiques EPFL - SB - ISIC
Institution of higher education EPF Lausanne - EPFL
Main discipline Biochemistry
Start/End 01.12.2019 - 30.11.2020
Approved amount 103'130.00
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All Disciplines (6)

Discipline
Biochemistry
Cellular Biology, Cytology
Organic Chemistry
Molecular Biology
Genetics
Pharmacology, Pharmacy

Keywords (5)

mechanistic chemical-biology tools development; reactive electrophilic lipid-meabolite signaling; split protein technology; interorganellar contactology; subcellular small-molecule signaling

Lay Summary (French)

Lead
JUMP
Lay summary

Une des caractéristiques la plus importante des cellules eucaryotes est la compartimentalisation. Dedans les cellules il y a des structures concrètes et indépendantes, qui s’appellent organites. Chacun de ces organites présent un environnement spécifique et fonctionne dans une façon particulaire. Les défaillances avec les organites sont associées avec les maladies. Par exemple, le cancer est particulièrement lié aux changements chez les mitochondries. Pendant plusieurs années il était cru que les organites aient essentiellement des fonctions indépendantes. En revanche, les données récentes racontent une histoire de la coopération; aujourd’hui beaucoup de rapports disent que les organites différents se touchent, formant des régions de chevauchement, qui sont importantes pour la santé. Malheureusement, ces régions de chevauchement sont transitoires et constituent beaucoup moins que la moitié de chaque organelle. Même avec les techniques de point, c’est vraiment difficile de les étudier.

Mais l’environnement hébergé par le chevauchement de organites est très diffèrent que l’environnement de chaque organite en isolation. Donc nous proposons que ces régions de chevauchement cacheraient protéines spécifiques ayant des propriétés très différentes que les protéines en vrac, qui existent soit dans les organites isolés soit même dans plusieurs autres endroits dans les cellules. Nous allons découvrir ces contexte-spécifique protéines utilisant une technique qui s’appelle «Jump» (sauter en Français). Cette méthode est une combinaison de deux techniques de pointe, fondées sur les aperçus de la biologie moléculaire. Avec «jump» nous pourrons se mater ces jonctions avec une loupe chemique, très puissante, qui peut ouvrir une nouvelle fenêtre pour les étudier et les maîtriser.

Direct link to Lay Summary Last update: 18.11.2019

Responsible applicant and co-applicants

Employees

Name Institute

Abstract

The eukaryotic cell is a hive of compartmentalized environments. These environments-such as the endoplasmic reticulum (ER) and the mitochondria-were long considered to be separated from each other. Mounting evidence now indicates that transient associations between numerous organelles forge highly-coordinated environments essential for cross-compartment information flow. Proteins that reside within these discrete sites likely experience transient changes to their interactomes and small-molecule-signal inducers during these associations. This is because at such junctions, organelles with intrinsically different properties, such as the mitochondria, a key source of reactive electrophilic species, and the ER, an organelle less associated with electrophile generation, come together. It is thus reasonable to posit that electrophilic signaling could readily be elevated at mitochondria-ER junctions, above that of non-electrophile-stimulated ER (or even mitochondria). We thus hypothesize that there are specific proteins within the ER and/or mitochondria that sense and respond to electrophilic signals, specifically at these junction interfaces. However, mitochondria-ER junctions comprise only 5-20% of the total surface area of these organelles. Thus, using traditional target-identification methods, proteins intrinsically electrophile sensitive at these junctions are lost in the noise of the bulk proteome. Emerging methods to probe junction-specific proteins have only the ability to define/identify proteins present at these junctions. Although mapping sub-organellar proteins is important to building up a picture of subcellular processes, it is even more critical to zero-in on proteins that have context-dependent gain-of-function signaling properties, since it is these proteins that likely marshal key regulatory events, and are often critical nexuses of major signaling cascades. Indeed, altered signal transduction processes are implicated in ER-mitochondria tethers. However, we have not even scratched the surface in terms of underpinning mechanisms. Unfortunately, no tools that can directly and precisely probe functions and signal responsivity within these interorganellar junctions are available to date.By adapting a recently-published target-identification method, G-REX, which has the potential to identify such unique and context-regulated interfacial events, this proposal will functionally deconstruct signaling architectures present in the hitherto murky areas of membrane junctions. By combining split-protein technology anchoring each of the two (N- vs. C-terminus-bearing) “split” components of HaloTag-protein to the outer membranes of the mitochondria and ER respectively, our proposed system, JUMP (Junction-specific mapping of interorganellar sensors and their signaling activities), selectively localizes small-molecule photocaged probes (precursors to specific lipid-derived electrophiles) to mitochondria-ER junctions. Uncaging allows controlled and transient delivery of the electrophile, such that lipid-sensitive proteins specifically at such junctions will be identified. T-REX, the established companion tool of G-REX, will then probe functional ramifications of covalent electrophile-binding of the specific hits we identify in the JUMP:REX screen. Understanding context-dependent signaling is of paramount importance to illuminate signaling events, map pathway intersections, and ultimately develop targeted interventions. However, despite the growing appreciation of links between defects in interorganellar contacts and neurodegenerative diseases, tools to probe or understand protein players, and importantly, their signaling activities/signal responsivity in the context of interorgenallar junctions remain inexistent. Thus, functional studies derived from JUMP can provide a lens through which to not only identify important proteins, but also glean new functional understanding.
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