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Dynamics of the Light Harvesting Complex network
Titre du projet
Dynamics of the Light Harvesting Complex network
Description
Efficient use of the light is essential for the fitness of photosynthetic organisms. It is believed that the evolution of an antenna network system, capable of dynamic regulation of the fraction of light energy to be transferred to the photosystems and to dissipate light excess is a key step in the evolution of photosynthetic eukaryotes and a crucial step in land colonization. The core structure of the pigment-protein complexes devoted to light harvesting (LHCs) is largely conserved among eukaryotes. However, the regulatory portions display a certain degree of variability which may underlie their role in specific adaptation to particular climatic conditions. The aim of the proposed research project is to understand the network of interactions between the proteins of the thylakoid membrane that allows coping with the changes in light quality and regime exploiting the model organisms Arabidopsis thaliana. In particular, the project will focus on the contribution of the N-terminal domain of the major light harvesting antenna (LHCII) and its phosphorylation to the regulatory dynamics of the thylakoids. The primary focus will be to understand the contribution of the two major isoforms of the LHCII complex LHCB1 and LHCB2. These being the major components of the trimeric LHCII and dynamically phosphorylated at the N-terminus. At the end of the proposed project the specific role of each isoforms’ N-terminus in vivo for the regulation of photosynthetic electron transfer, the assembly of photosynthetic supercomplexes and the architecture of the thylakoid membrane network will be revealed. The key tool for the investigation will be the production of Arabidopsis lines expressing mutated versions of the major light harvesting complex isoforms. These will be obtained by complementing previously produced knock-out lines with LHCII isoforms containing a modified phosphorylation site. By analyzing their photosynthetic performance and photosystems structure under changing light regimes it will be possible to elucidate the role of their dynamic interaction with other components of the photosynthetic machinery. Furthermore, the results will allow the elaboration of hypotheses on the evolution of this regulation system. Moreover, since antenna re-organization is central in the photosynthetic acclimation responses, the new detailed knowledge should allow informed manipulations of its kinetics. It has already been demonstrated that by reducing the response time it is possible to engineer a higher productivity in the field. Furthermore, understanding these mechanisms will offer new targets for genetic selection in crop breeding programs and provide concepts applicable to the design of artificial-photosynthesis systems.
Chercheur principal
Statut
Ongoing
Date de début
1 Septembre 2018
Date de fin
1 Septembre 2022
Chercheurs
Pagliano, Cristina
Finazzi, Giovanni
Croce, Roberta
Organisations
Identifiant interne
40906
identifiant
1 Résultats
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- PublicationAccès libreEngineering Light Harvesting Complex IITo sustain the life on Earth solar energy has to be converted into chemical energy, this is possible thanks to the process called photosynthesis. Through a set of interconnected pigment-binding proteins, collectively called “light harvesting complex” (LHC), photon energy is collected and funneled towards the reaction centers (RCs) of the two photosystems. The RCs feed a series of redox reactions that ultimately allow the production of reducing power (NADPH) and ATP, used for the synthesis of organic molecules. Despite having a highly conserved core structure, the light harvesting complex II (LHCII) is capable to acclimate to a wide range of environmental conditions. LHCII can functionally associate to both photosystems thus allowing a fine tuning of the electron transport chain, or act as dissipators of excess light energy protecting the photosystems from photodamage. This dynamic regulation is crucial for the adaptation of plants to different light conditions. LHCII is mostly composed of homo and hetero trimers of three isoforms: LHCB1, LHCB2 and LHCB3. The first two, through the phosphorylation of a threonine present in the N-terminal domain, are crucial for the regulation of the dynamics of the LHCII network. LHCB2 phosphorylation plays a central role in LHCII association to photosystem I and is thus regarded as the regulatory isoform of LHCII. The role of LHCB1 phosphorylation is less obvious; however, it has an impact in vivo allowing a partial adaptive response also in absence of LHCB2. Thanks to the CRISPR/Cas9 technique, we produced multiple mutants for the clustered genes coding for LHCB1 and LHCB2, thus allowing the production of complete null mutants for these two LHCII isoforms. These mutant lines constitute an ideal platform to study the impact of targeted modifications on the LHCII network via the production of complemented lines. LHCB1 is the most abundant isoform of LHCII and, consequently, a multiple mutation of the five genes encoding this protein results in a pale phenotype, reduced PSII antenna cross-section, altered thylakoid structure along with lower Photosystem I over Photosystem II reaction center ratio. Interestingly, the loss of one of these two major isoforms results in compensatory effects at the phosphorylation level of the remaining. Loss of LHCB1 results in a de-phosphorylation of the remaining LHCB2, while loss of LHCB2 results in an over-phosphorylation of LHCB1. The complete knock out plants for both LHCB1 and LHCB2 were tested under prolonged fluctuating light, moderate temperature stress and their combination. This revealed an increased susceptibility to only the combined stress for the complete LHCB1 knock out, visible as a clear growth delay, combined with a decrease in the photosynthetic efficiency. Surprisingly, the loss of LHCB2, which impairs the antenna re-allocation between the two photosystems, did not result in any major defect under the combined stress condition. Modification of the threonine of the phosphorylation site to alanine (non phosphorylable) or aspartate (constitutive negative charge "phospho-mimic») for both LHCB1 and LHCB2 reveals the impact of such irreversible modification on photosynthetic acclimation and on the dynamics of the photosynthetic complexes. We demonstrated that the complete removal of LHCB2 protein or the substitution of its phosphorylation site by alanine or aspartate largely, result in a physiologically overlapping phenotype with sharp reduction of state transitions and decreased LHCII-PSI-LHCI supercomplex formation. These defects result in slower acclimation to fluctuating light. Our results show that only with the phospho-threonine group LHCB2 can fully accomplish state transitions and that the negative charge of the aspartate substitution has no impact on short-term photosynthetic acclimation. Disentangling the defined role of each antenna isoform, LHCB1 and LHCB2, could shed light in short and long term acclimatory processes. Leading to a better comprehension on how each isoform contributes to LHCII network organization and results in an optimal balance between light capture and photoprotection. The multiple null lines produced during this project are a milestone along this path and open future perspectives towards the design of innovative LHCII complementation studies.