Dual urea utilization enzyme systems in Symbiodiniaceae coral symbionts under warming

Urea has been shown to be important as a nitrogen (N) nutrient for coral holobionts, but the mechanism underpinning urea utilization by symbiotic algae is not fully understood. In this study, we investigated the molecular pathways underlying urea utilization in the Symbiodiniaceae family and the responses of these pathways to different N-nutrient conditions and heat stress through comprehensive genomic screening, multi-omics analysis and stable isotope pulse-chase experiments. Genome screening revealed that two urea hydrolysis systems, urease (URE) and urea amidolyase (UAL), were present in Symbiodiniaceae, positioning this lineage as one of the few non-green algae that possess UAL. Furthermore, our data reveal an interesting evolutionary trajectory of UAL. While subunit DUR2 occurs in most symbiodiniacean genomes sequenced to date, only two species (Cladocopium goreaui and Cladopium c92) possess the complete UAL system (DUR1 with DUR2). In the phylogenetic tree of UAL sequences, Symbiodiniaceae clustered more closely with coral symbiotic bacteria than with other eukaryotes, but show clear distinct genetic features such as GC content and codon usage, suggesting evolutionary horizontal gene transfer from bacteria. Furthermore, ex-hospite C. goreaui exhibited better growth and achieved higher maximum specific growth rates when urea was provided as the sole nitrogen source, compared to ammonium. Notably, when experimenting on the Cladocopium-dominating Pocillopora damicornis holobiont using 15N isotope tracer, we found that under heat stress (HS) conditions, the in-hospite Symbiodiniaceae significantly increased urea uptake but decreased NO3− and NH4+ uptake. Omics analyses suggest that responses to different nitrogen, light, and temperature conditions were more likely mediated by UAL. This study reveals two distinct urea utilization systems in the coral ecosystem and their differential responses to warming, highlighting the importance of urea as N-nutrient when facing global warming.