Characterization of photosystem I assembly and photoprotection in Hordeum vulgare and Physcomitrium patens

Oxygenic photosynthesis is a biochemical process that converts solar light energy into chemical energy, fixing carbon dioxide (CO2) into organic molecules and producing molecular oxygen (O2) as a byproduct. In eukaryotic photosynthetic organisms, this process occurs in the chloroplast, an organelle that contains a system of membranes known as thylakoids, where two light-absorbing complexes, photosystem I (PSI) and photosystem II (PSII), are located. These complexes, whose biogenesis involves a network of highly conserved assembly factors and proteins, contain chlorophyll a molecules that undergo charge separation and trigger electron transport when excited by light energy. The preeminent form of photosynthetic electron transport is linear electron transport, where electrons flow from PSII to the transmembrane cytochrome b6f complex, the soluble plastocyanin, PSI, and the stromal ferredoxin, in this order, to ultimately reduce NADP+, generating the reducing agent NADPH used in CO2 fixation. Nonetheless, under stressful light conditions, such as sudden dark-to-light transitions and high light intensities, this process is coupled with the production of reactive oxygen species (ROS), which can significantly disrupt the photosynthetic apparatus’s efficiency. To counter these issues, several protective mechanisms have evolved in photosynthetic organisms, such as alternative forms of electron transport, which avoid the over-reduction of electron acceptors and decrease ROS formation. One of these alternative electron transport pathways is the pseudo-cyclic electron transport, which involves the ferredoxin-mediated reduction of O2. This process can be induced by the spontaneous Mehler reaction, generating the radical superoxide (O2•−), which is then converted into water by the sequential activity of superoxide dismutases (SOD) and ascorbate peroxidases. One of the aims of this thesis was the characterization of the Hordeum vulgare cv. Morex TM-1214 mutant, which harbours a single amino acid substitution (S362F) in the High Chlorophyll Fluorescence 101 (HCF101) protein, a highly conserved scaffold protein that is crucial for the assembly of PSI. The functionality of the photosynthetic machinery in this mutant was assessed using spectrometric techniques, revealing several alterations of the photosynthetic apparatus, such as increased energy dissipation as heat in the form of non-photochemical quenching and a decrease in the content of functional PSI. Differences were also detected in CO2 fixation, which appeared to be impaired in TM-1214 when compared to the wild type. The second aim of this thesis was the characterization of the two iron superoxide dismutases (FeSOD-A and FeSOD-B) present in Physcomitrium patens. To do this, P. patens reporter lines encoding FeSODs fused with the Yellow Fluorescent Protein (YFP) were generated and isolated. The effects of the expression of these fused proteins were assessed through growth tests conducted by exposing plants to different growth light conditions, revealing no significant differences in their growth and photosynthetic performance compared to the wild type.