Heterologous expression of genes from a heterocystous cyanobacterial endosymbiont highlights organic carbon exchange with its diatom host

ABSTRACT A few genera of diatoms are widespread and thrive in low nutrient waters of the open ocean due to their close association with N 2 -fixing, filamentous heterocyst-forming cyanobacteria. In one of these symbioses, the symbiont, Richelia euintracellularis , has penetrated the cell envelope of the host, Hemiaulus hauckii , and lives inside the host cytoplasm. How the partners interact, including how the symbiont sustains high rates of N 2 fixation is unstudied. Since R. euintracellularis has evaded isolation, heterologous expression of genes in model laboratory organisms was performed to identify the function of proteins from the endosymbiont. Gene complementation of a cyanobacterial invertase mutant and expression of the protein in Escherichia coli showed that R. euintracellularis HH01 possesses a neutral invertase that splits sucrose producing glucose and fructose. Several solute binding proteins (SBPs) of ABC transporters encoded in the genome of R. euintracellularis HH01 were expressed in E. coli and their substrates were characterized. The selected SBPs directly linked the host as the source of several substrates, e.g., sugars (sucrose, galactose), amino acids (glutamate, phenylalanine) and a polyamine (spermidine), to support the cyanobacterial symbiont. Finally, transcripts of genes encoding the invertase and SBPs were consistently detected in wild populations of H. hauckii collected from multiple stations and depths in the western tropical North Atlantic. Our results support the idea that the diatom host provides the endosymbiotic cyanobacterium with organic carbon to fuel N 2 fixation. This knowledge is key to understand the physiology of the globally significant H. hauckii-R. euintracellularis symbiosis. SIGNIFICANCE Diatom diazotroph associations (DDAs) between diatoms and N 2 -fixing bacteria (diazotrophs) have a relevant impact on N 2 fixation-based production, but the mechanisms underlying their integrated N 2 and CO 2 fixation remain unstudied. In the association between the diatom Hemiaulus hauckii (host) and the N 2 -fixing, heterocyst-forming cyanobacterium Richelia euintracellularis (endosymbiont), the cyanobacterium is uncultivable. Here we used heterologous expression of genes from the endosymbiont to identify the function of proteins involved in the utilization of organic carbon from the host. The importance of these proteins was also confirmed by estimating gene expression in environmental samples. Our results show that the metabolisms of the symbiotic partners are integrated allowing the host to sustain the physiology of the endosymbiont for an important ecological role.