Nitrate assimilation in the coral-algal symbiosis : understanding the patterns of expression of the enzyme nitrate reductase in Symbiodiniaceae.

Coral holobionts consist of complex associations between a coral host, microalgae (Symbiodiniaceae) and a microbiome. The oligotrophic nature of coral reef environments implies that corals rely heavily on the nutrient exchanges with their photosynthetic endosymbionts to function. In addition to providing translocated photosynthates, the algae account for most of the uptake of dissolved inorganic nitrogen. Coral hosts are unable to reduce nitrate as they lack the necessary enzymes, whereas Symbiodiniaceae have been shown to express the enzyme nitrate reductase (NR). However, the evidence supporting the active reduction of nitrate by the symbiotic algae during symbiosis is scarce and equivocal. Rigorous studies on nitrate assimilation by Symbiodiniaceae are lacking yet are essential for understanding coral holobiont functioning. Using western-blotting and qRT-PCR, we studied the expression of the enzyme nitrate reductase in free-living Symbiodiniaceae under varying conditions to decipher the mechanisms underlying its regulation. We show that the expression and regulation of NR is a dynamic and reversible process impacted by NO3- and NH4+ concentrations. In particular, we demonstrate that NR is actively degraded in the presence of NH4+ and that the synthesis of NR is light-dependent. Additionally, qRT-PCR assays suggest that NR expression is regulated at the post-transcriptional stage as the NR-coding gene is expressed uniformly across conditions. Because NR has never been identified in in hospite coral symbionts, we next investigated the expression of NR in Symbiodiniaceae during symbiosis. For this purpose, different coral species were depleted in nitrogen before they were exposed to an enrichment in NO3-. We demonstrate, for the first time, that Symbiodiniaceae express NR during symbiosis, and that this expression is dependent on NO3- concentrations. This study deciphers the functioning of an essential enzyme involved in coral nutrition, shedding a new light and raising new questions on the process of inorganic nitrogen assimilation by coral holobionts.