The cells in general, and its subcellular compartments mitochondria and chloroplasts in particular, have a high iron demand to ensure the functionality of vital processes such as photosynthesis, respiration, sulfur and nitrogen assimilation and chlorophyll metabolism. All these metabolic pathways require the presence of numerous metalloproteins including those containing iron-sulfur (Fe-S) clusters or hemes. Similarly, other processes such as DNA repair and replication, ribosome biogenesis, tRNA thio-modification or co-enzyme (biotin, lipoic acid, thiamine) synthesis also rely on the functionality of Fe-S proteins (Lill, 2009). In both eukaryotes and prokaryotes, Fe-S proteins are first synthesized as apoforms before the post-translational insertion of the cofactor into the polypeptides through specific assembly machineries found in the cytosol (CIA), in mitochondria (ISC) and in chloroplasts (SUF) (Couturier et al., 2013). The incorporation of Fe-S clusters into proteins is classically described as a two-step process. The first step is the de novo assembly of Fe-S clusters onto scaffold proteins and the second step is the transfer of the preformed clusters to acceptor proteins through the action of transfer proteins.
The general objective of this PhD proposal is to understand the functioning of the Fe-S cluster assembly machineries in organelles and more precisely the molecular mechanisms controlling the second step, i.e. the trafficking of Fe-S clusters from scaffold proteins to final acceptors. This project will mainly focus on two insufficiently characterized protein families (NFU and SUFA/ISCA) assumed to participate in the trafficking of Fe-S clusters associated with the plastidial or mitochondrial machineries. In collaboration with national and international partners, we are currently developing a multidisciplinary approach combining plant genetics and physiology, molecular and structural biology and biochemistry approaches. More precisely, in order to delineate the exact roles of NFU and SUFA/ISCA proteins in the Fe-S cluster machineries present in plant organelles, we would like to address the following central questions:
(i) Do these proteins assemble different types of Fe-S clusters and do they have specific interaction partners?
Based on the idea that some proteins could display alternate biochemical functions, integrate different types of cluster or interact with different substrate proteins in different physiological situations, as shown for Escherichia coli proteins (Roche et al., 2013), we will perform analytical, biochemical and spectroscopic analyses of anaerobically-purified recombinant proteins produced in E. coli. Moreover, we will use an unprecedented variety of approaches (pull-down, co-immunoprecipitation, and yeast two hybrid assays) to define the interaction network of each candidate.
(ii) What are the physiological consequences of deleting the genes and is the function conserved across kingdoms?
The physiological and phenotypic analysis of loss-of-function or overexpressing plant lines will be undertaken under various regimes of photoperiod, temperature and light intensity as well as under specific environmental constraints (iron and sulfur starvation or excess). Moreover, complementation experiments of yeast mutants will be performed to understand whether the function is evolutionary conserved but also simply to assess the roles of the proteins when A. thaliana insertion mutants are lethal or just not available.
Overall, understanding the mechanisms of Fe-S cluster assembly and trafficking is crucial considering that plants contain several dozens of Fe-S proteins, many of which are essential for normal cellular metabolism and energy supply. This is well illustrated by the fact that the disruption of many genes coding for components of these biosynthetic machineries in plants, mammals and yeast leads to severe disorders/diseases (Lill, 2009). Furthermore, the chloroplast compartment is unique to photosynthetic organisms, and it is fully autonomous for its Fe-S synthesis. Hence, this process is essential for the photosynthetic activity which sustains an optimal plant growth.
The PhD work will primarily be carried out in the UMR 1136 UL-INRA “Tree-Microbe interactions” in the team “Stress response and redox regulation” but mobility will be necessary for exchange periods with national and international collaborators.
Pr Nicolas Rouhier – UMR Tree-Microbe Interactions (IAM)
Faculté des sciences et technologies, F-54500 Vandoeuvre-lès-Nancy
How to apply
In order to prepare a PhD thesis within the Lorraine Université d’Excellence Program, the interested candidate should consult the PhD topics offered in each social and economic challenges.
These PhD thesis topics are proposed by faculty members or researchers accredited to supervise research.
Candidate application period: according to graduate school schedule (visit each topic)
Each candidate may submit an application on up to three separate research topics.
Application analysis period by each graduate school
The graduate school reviews the applicants for a doctoral contract in the relevant disciplines. They check the level of supervision for each supervisor and the situation of trained doctors. Each candidate will meet the laboratory director, supervisor and a representative from the graduate school. This interview is to identify the candidate’s motivations and suitability as a candidate for the PhD project proposed by the supervisor. A recommendation will be made to the graduate school. This will summarize the strengths and/or weaknesses of the application.
PhD grants will include monthly income for the PhD student (roughly 1700 € for research only, complement can be provided for teaching missions) and environment for research in the research unit.
Please be aware that in order to offer a variety of subjects, more positions are posted here than available funding. The LUE executive committee will make the final choice on the granted funding (up to 12 positions), based on the recommendations by the doctoral schools.