Interplay between non-photochemical plastoquinone reduction and re-oxidation in pre-illuminated Chlamydomonas reinhardtii: a chlorophyll fluorescence study

In photosynthetic eukaryotes, the redox state of the plastoquinone (PQ) pool is an important sensor for mechanisms that regulate the photosynthetic electron transport. In higher plants, a multimeric nicotinamide adenine dinucleotide (phosphate) (NAD(P))H dehydroge- nase (NDH) complex and a plastid terminal oxidase (PTOX) are involved in PQ redox homeostasis in the dark. We recently demonstrated that in the microalgae Chla- mydomonas reinhardtii, which lacks the multimeric NDH complex of higher plants, non-photochemical PQ reduction is mediated by a monomeric type-II NDH (Nda2). In this study, we further explore the nature and the importance of non-photochemical PQ reduction and oxidation in relation to redox homeostasis in this alga by recording the ‘dark’ chlorophyll fluorescence transients of pre-illuminated algal samples. From the observation that this fluorescence tran- sient is modified by addition of propyl gallate, a known inhibitor of PTOX, and in a Nda2-deficient strain we conclude that it reflects post-illumination changes in the redox state of PQ resulting from simultaneous PTOX and Nda2 activity. We show that the post-illumination fluo- rescence transient can be used to monitor changes in the relative rates of the non-photochemical PQ reduction and reoxidation in response to different physiological situa- tions. We study this fluorescence transient in algae acclimated to high light and in a mutant deficient in mitochondrial respiration. Some of our observations indi- cate that the chlororespiratory pathway participates in redox homeostasis in C. reinhardtii

Function of the chloroplastic NADP(H) dehydrogenase NDA2 for the H2 photoproduction in sulphur-deprived Chlamydomonas reinhardtii

The relative contributions of the PSII-dependent and Nda2-dependent pathways for H2 photoproduction were investigated in the green microalga Chlamydomonas reinhardtii after suphur-deprivation. For this purpose, H2 gas production was compared for wild-type and Nda2-deficient cells with or without DCMU (a PSII-inhibitor) in the same experimental conditions. Nda2-deficiency caused a 30 % decrease of the maximal H2 photoevolution rate observed shortly after the establishment of anoxia, and an acceleration of the decline of H2 photoevolution rate with time. DCMU addition to Nda2-deficient cells completely inhibited H2 photoproduction, showing that the PSII-independent H2 photoproduction relies on the presence of Nda2, which feeds the photosynthetic electron transport chain with electrons derived from oxidative catabolism. Nda2-protein abundance increased as a result of sulphur deprivation and further during the H2 photoproduction process, resulting in high rates of non-photochemical plastoquinone reduction in control cells. Nda2-deficiency had no significant effect on photosynthetic and respiratory capacities in sulphur-deprived cells, but caused changes in the cell energetic status (ATP and NADPH/NADP+ ratio). The rapid decline of H2 photoevolution rate with time in Nda2-deficient cells revealed a more pronounced inhibition of H2 photoproduction by accumulated H2 in the absence of non-photochemical plastoquinone reduction. Nda2 is therefore important for linking H2 photoproduction with catabolism of storage carbon compounds, and seems also involved in regulating the redox poise of the photosynthetic electron transport chain during H2 photoproduction.ARC: Micro-H

A novel force sensor with zero stiffness at contact transition based on optical line generation

International audienceRobotization of medical acts often requires the evaluation of contacts between a robotic system and a patient, for safety or efficiency reasons. When contact occurs with a stiff environment, instabilities and vibrations can appear and a passive compliance is therefore needed. In this paper, we propose to embed compliance in a force sensor and to develop a novel force sensor with large compliance, i.e. a zero stiffness at contact transition to ease robot control. To get at the same time a satisfying measurement range and low off-axis sensitivity, an optical measurement process based on an optical line generated thanks to additive manufacturing is exploited. A compliant sensor body allowing the desired stiffness profile is presented and the specific optical measurement technique is developed. Finally, a prototype of the proposed force sensor is evaluated experimentally

Analyse fonctionnelle de la production d'hydrogène chez des mutants mitochondrieux de Chlamydomonas reinhardtii

Mitochondrial Chlamydomonas mutants for respiratory complexes present a decreased dark respiration and apparent yield of photosynthetic linear electron flow. They accumulate reducing power such as NAD(P)H and show lower levels of ATP. Under restrictive conditions, like sulfur depletion and anoxia, Chlamydomonas is able to produce hydrogen towards the activation of a chloroplatic O2-sensitive Fe-hydrogenase which catalyses the reduction of electrons to H2. In this study we used an adapted Melis protocol to analyse hydrogen evolution of mitochondrial mutants. For this aim a simple-flask system was built with gaz collecting tubes. A parallel flask was used for GC analyses

Expression of recombinant enhanced green fluorescent protein provides insight into foreign gene-expression differences between Mut+ and MutS strains of Pichia pastoris.

peer reviewedPichia pastoris is a very popular yeast for recombinant protein production, mainly due to the strong, methanol-inducible P(AOX1) promoter. Methanol induction however poses several drawbacks. One approach to improve processes is to use MutS strains with reduced methanol catabolic ability. Various reports claim that MutS allows higher recombinant protein production levels than Mut+ but scarcely elaborate on reasons for differences. In this study, enhanced green fluorescent protein was used as a P(AOX1) -driven reporter for the investigation of expression differences between Mut+ and MutS strains. Mut+ exhibited higher responses to methanol, with faster growth (0.07 vs. 0.01 hr(-1) ) and higher consumption of methanol (2.25 vs. 1.81 mmol/g(DCW) .hr) and oxygen (2.2 vs. 0.66 mmol/g(DCW) .hr) than MutS. Mut+ yielded more biomass than MutS (2.3 vs. 1.3 g(DCW) /L), and carbon dioxide analysis of bioreactor off-gas suggested that considerable amounts of methanol were consumed by Mut+ via the dissimilatory pathway. In contrast, it was demonstrated that the MutS population switched to an induced state more rapidly than Mut+. In addition, MutS exhibited 3.4-fold higher fluorescence levels per cell (77,509 vs. 23,783 SFU) indicative of higher recombinant protein production. The findings were verified by similar results obtained during the expression of a lipase. Based on the differences in response to methanol versus recombinant protein production, it was proposed that higher energy availability occurs in MutS for recombinant protein synthesis, contrary to Mut+ that uses the energy to maintain high levels of methanol catabolic pathways and biomass production