RNA-protein dynamics of high CO2-adapted, iron-phosphorus co-limited Trichodesmium reveals nutrient-limited molecular architecture of a globally significant marine nitrogen fixer in a future ocean

Our paper presents the results of a study in which we used Illumina RNA-Seq (i.e. transcriptomics) and high-CO2 nutrient limitation experiments to examine transcriptional variation of iron-limited, phosphorus-limited, and iron-phosphorus co-limited cultures following long-term (~7 years) low- (380 µatm CO2) and high-CO2 (750 µatm CO2) selection. Hence, we describe the molecular physiology of the globally-significant, biogeochemically-critical marine cyanobacterium Trichodesmium. A single IMS101 population, originally isolated from an individual Trichodesmium colony, was split into low (380-selected) and high (750-selected) CO2 treatments of 6 biological replicates each, and grown for ~7 years (~1,000 and ~1,500 generations for 380- and 750-selected, respectively). Semi-continuous culturing methods were used with a light-dark cycle of 12:12 (light:dark) at 26 C in a modified Aquil medium (devoid of fixed nitrogen) containing standard vitamins and trace metals with 250 nM Fe and 10 µM phosphate (PO43-). Next, Fe/P co-limited treatments were generated by sub-culturing 3 biological replicates from each CO2 treatment and incubating them in the aforementioned media except with co-limiting levels of Fe (10 nM) and PO43- (0.25 µM ) for >1 year. To then mimic episodic pulses of Fe or P experienced by Fe/P co-limited microbes in the oligotrophic ocean , either Fe or P was added at replete concentrations to subcultures of the Fe/P co-limited cell lines to generate P and Fe limited lines at both CO2 levels, respectively. Following a two-month acclimation period to create steady-state conditions, the lines were sampled for molecular analyses. All samples for transcriptomics (biological duplicates) were taken by filtering cells onto 5-µm polycarbonate filters during the middle of the photoperiod, flash frozen, and stored in liquid nitrogen until extraction.