PhD position

Zinc-induced protein aggregation in neurodegenerative disease: TPD-43 and Amyotrophic Lateral Sclerosis (ALS)

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In case of success, we will be able to propose new pathological pathway of ALS development and new drug targets for ALS therapy.

Description

The PhD project below is part of NeuroSchool’s “PhD Scholarships for International Students” call, which will fund 3 scholarships among the nine proposed projects.
APPLICATION MUST PROCEED THROUGH OUR WEBSITE: https://neuro-marseille.org/en/Calls/phd-scholarships-for-international-students/

STATE OF THE ART : Most neurodegenerative diseases are characterized by the presence of aggregation-prone proteins associated with the pathology. For instance, amyloid- β and tau are found aggregated in Alzheimer’s disease, α-synuclein in Parkinson disease, TDP-43, FUS and SOD1 in ALS. Remarkably, these proteins, which differ so much in their structures and functions, share common features. For instance, most of them are known to bind zinc ions. While zinc might not be the causative agent of these diseases, it was shown to bind to these proteins and favor their aggregation. Until recently, TDP-43 was not recognized as a zinc-binding one. It does not contain any known zinc-binding motifs. Still, recently together with our colleagues from Moscow State University we have demonstrated that TDP-43 is able to binds zinc and showed that this binding significantly decreases stability and induces its irreversible aggregation (Garnier et al. 2017).

OBJECTIVES : (1) Identify TDP-43 zinc-binding site and resolve its 3D structure; (2) Determine the role of this site in zincinduced aggregation (in vitro, in cell and in vivo)

METHODS : To achieve these objectives, the application of a large number of methods both in vitro and in vivo is necessary. In the frame of the first objective, a set of TDP-43 mutants and fragments will be generated using standard biochemical approaches (direct mutagenesis, HPLC etc). Then interaction of purified mutants with zinc will be studied using biophysical methods such as Isothermal Titration Calorimetry (ITC), Differential scanning Calorimetry and fluorimetry (DSC and nanoDSF). Finally, the complex of TDP-43 with zinc will be characterized using structural methods, such as NMR. To achieve the second objective the aggregation of TDP-43 mutants and its fragments will be studied in vitro using Turbidimetry and Dynamic Light Scattering (DLS) and in cells using GFP constructs, microscopy, flow-cytometry. At last, TDP- 43 aggregates from brain of ALS animal models will be purified to measure the content of metal ions, in particularly zinc.

EXPECTED RESULTS : We expect to determine the amino acids that chelate zinc ions in TDP-43 and resolve its 3D structure in complex with zinc. By mutating these amino acids, we expect to reduce significantly zinc-induced aggregation of TDP-43. Also, we expect to find zinc in TDP-43 aggregates extracted from the brain of ALS animal models. In case of success, we will be able to propose new pathological pathway of ALS development and new drug targets for ALS therapy.

FEASABILITY : Institute of NeuroPhysiopathology and collaborators have all necessary equipment and infrastructure to accomplish proposed research project. Moreover, similar project on tau zinc-induced aggregation have been successfully performed in the laboratory by another PhD student.

Desired profile

We are looking for a motivated candidate with good fundamental background and who is willing to acquire complementary expertise in in vitro, in cell and in vivo approaches.

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