Impaired photoprotection in Phaeodactylum tricornutum KEA3 mutants reveals the proton regulatory circuit of diatoms light acclimation

International audienceDiatoms are successful phytoplankton clades able to acclimate to changing environmental conditions, including e.g. variable light intensity. Diatoms are outstanding at dissipating light energy exceeding the maximum photosynthetic electron transfer (PET) capacity via the nonphotochemical quenching (NPQ) process. While the molecular effectors of NPQ as well as the involvement of the proton motive force (PMF) in its regulation are known, the regulators of the PET/PMF relationship remain unidentified in diatoms. We generated mutants of the H$^+$ /K$^+$ antiporter KEA3 in the model diatom $Phaeodactylum\ tricornutum$. Loss of KEA3 activity affects the PET/PMF coupling and NPQ responses at the onset of illumination, during transients and in steady-state conditions. Thus, this antiporter is a main regulator of the PET/PMF coupling. Consistent with this conclusion, a parsimonious model including only two free components, KEA3 and the diadinoxanthin de-epoxidase, describes most of the feedback loops between PET and NPQ. This simple regulatory system allows for efficient responses to fast (minutes) or slow (e.g. diel) changes in light environment, thanks to the presence of a regulatory calcium ion (Ca$^{2+}$ )-binding domain in KEA3 modulating its activity. This circuit is likely tuned by the NPQ-effector proteins, LHCXs, providing diatoms with the required flexibility to thrive in different ocean provinces

Mapping plastid transcript population by circular reverse transcription polymerase chain reaction

International audienceDuring evolution of photosynthetic organisms, the genetic information provided by the internalized cyanobacteria has been transferred to the nucleus. The small genome kept by the chloroplast, the so-called plastome, displays a complex organization, comprising operons under the control of multiples promoters. In addition, polycistronic transcripts undergo multiple processing events, thus generating a complex population of mRNAs from a single gene. This chapter describes a method to investigate the diversity of the mRNA population from a single gene by circular RT-PCR. The protocol provided here allows for the simultaneous mapping of both 5' and 3' ends of the same RNA molecule

An update on the regulation of photosynthesis by thylakoid ion channels and transporters in Arabidopsis.

In natural, variable environments, plants rapidly adjust photosynthesis for optimal balance between light absorption and utilization. There is increasing evidence suggesting that ion fluxes across the chloroplast thylakoid membrane play an important role in this regulation by affecting the proton motive force and consequently photosynthesis and thylakoid membrane ultrastructure. This article presents an update on the thylakoid ion channels and transporters characterized in Arabidopsis thaliana as being involved in these processes, as well as an outlook at the evolutionary conservation of their functions in other photosynthetic organisms. This is a contribution to shed light on the thylakoid network of ion fluxes and how they help plants to adjust photosynthesis in variable light environments

Thylakoid potassium channel is required for efficient photosynthesis in cyanobacteria

A potassium channel (SynK) of the cyanobacterium Synechocystis sp. PCC 6803, a photoheterotrophic model organism for the study of photosynthesis, has been recently identified and demonstrated to function as a potassium selective channel when expressed in a heterologous system and to be located predominantly to the thylakoid membrane in cyanobacteria. To study its physiological role, a SynK-less knockout mutant was generated and characterized. Fluorimetric experiments indicated that SynK-less cyanobacteria cannot build up a proton gradient as efficiently as WT organisms, suggesting that SynK might be involved in the regulation of the electric component of the proton motive force. Accordingly, measurements of flash-induced cytochrome b(6)f turnover and respiration pointed to a reduced generation of ΔpH and to an altered linear electron transport in mutant cells. The lack of the channel did not cause an altered membrane organization, but decreased growth and modified the photosystem II/photosystem I ratio at high light intensities because of enhanced photosensitivity. These data shed light on the function of a prokaryotic potassium channel and reports evidence, by means of a genetic approach, on the requirement of a thylakoid ion channel for optimal photosynthesis

A Toolkit for the Characterization of the Photoprotective Capacity of Green Algae

International audienceWhile light is a crucial energy source in photosynthetic organisms, if its intensity exceeds their photosynthetic capacity it may cause light-induced damage. A dominant photoprotective mechanism in plants and algae is the qE (quenching of energy), the major component of nonphotochemical quenching (NPQ). qE is a process that dissipates absorbed excitation energy as heat, ensuring cell survival even under adverse conditions. The present protocol gathers together a set of experimental approaches (in vivo chlorophyll fluorescence, western blotting, growth and cellular chlorophyll content at very strong light) that collectively allow for the characterization of the qE capacity of the model green algae Chlamydomonas reinhardtii