Abstract
Nitrogen (N) scarcity is a frequently encountered situation that constrains global biomass productivity. In response to N deficiency, cell division stops and photosynthetic electron transfer is downregulated, while carbon storage is enhanced. However, the molecular mechanism downregulating photosynthesis during N deficiency and its relationship with carbon storage are not fully understood. The Proton Gradient Regulator-like 1 (PGRL1) controlling cyclic electron flow (CEF) and Flavodiiron proteins involved in pseudo-(CEF) are major players in the acclimation of photosynthesis. To determine the role of PGRL1 or FLV in photosynthesis under N deficiency, we measured photosynthetic electron transfer, oxygen gas exchange and carbon storage in Chlamydomonas pgrl1 and flvB knockout mutants. Under N deficiency, pgrl1 maintains higher net photosynthesis and O2 photoreduction rates, while flvB shows a similar response compared to control strains. Cytochrome b6f and PSI are maintained at a higher abundance in pgrl1. The photosynthetic activity of flvB and pgrl1 flvB double mutants decreases in response to N deficiency similar to the control strains. Furthermore, the preservation of photosynthetic activity in pgrl1 is accompanied by an increased accumulation of triacylglycerol depending on the genetic background. Taken together, our results suggest that in the absence of PGRL1-controlled CEF, FLV-mediated PCEF maintains net photosynthesis at a high level and that CEF and PCEF play antagonistic roles during N deficiency. It further illustrates how nutrient status and genetic makeup of a strain can affect the regulation of photosynthetic energy conversion in relation to carbon storage and provides new strategies for improving lipid productivity in algae.
Significance statement Nitrogen (N) deficiency, an often-encountered phenomenon in nature, results in growth arrest, downregulation of photosynthesis and massive carbon storage in microalgae. However, more mechanistic insights involved in tuning photosynthetic electron transfer during N deficiency are required. Here, we provide evidence that a well-conserved protein in chlorophytes, the Proton Gradient Regulator-like 1 (PGRL1), is a key regulator of photosynthesis during N deficiency. In its absence, cells exhibited sustained photosynthesis thanks to the Flavodiiron (FLV) proteins. We propose that both PGRL1 and FLV, by having antagonistic roles in N deficiency, manage the redox landscape, carbon storage and biomass production. Our work revolves around the current paradigm of photosynthesis regulation during N deficiency and provides a new framework for improving biomass production and carbon storage in microalgae for biotechnological purposes.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
Competing Interest Statement: There are no conflicts of interest.
To consolidate our protein quantification, we have repeated all the immunoblots in 3 independent biological replicates for all strains. We have also performed the same set of immunoblots for the flvB and pgrl1 flvB mutants for all the time points (see Fig. 4 B and C). Now, the heatmap is replaced by a bar graph showing three independent values for each protein. Additionally, we have performed spectroscopic measurements in order to determine how N-deficiency impacts the acceptor/donor side limitation of PSI in the pgrl1 mutants. To make the paper easier to follow, detailed information about strains used in this work is now summarized in Supplemental Table 1 and Figure S1. To strengthen our conclusions concerning the link between the observed phenotype and the pgrl1 mutation we have generated another allele of pgrl1 in a different genetic background (CC125) using CRISPR-Cas9. This additional pgrl1CC125 mutant showed a similar phenotype as compared to the pgrl1-137AH regarding photosynthetic properties, but a different behaviour for lipid accumulation. As mentioned by the reviewer, lipid accumulation should be considered as a metabolic consequence rather than directly linked to PGRL1 function, and may therefore be affected by the genetic context. We have added a discussion section to address this issue. All the information regarding the CRISPR-Cas9 generated mutant is reported in Supplemental Figures 3, 4, 6 and Supplemental Tables 2 and 3.
