Here, we examined two-way and multiple driver effects of ocean acidification and other key environmental drivers - nitrate, phosphate, irradiance, and temperature - on the growth, photosynthetic and calcification rates, and the elemental composition of E. huxleyi. In addition, changes in functional gene expression were examined to understand the molecular mechanisms underpinning the physiological responses. The single driver manipulation experiments sugguest decreased nitrate supply being the most important driver regulating E. huxleyi physiology, by significantly reducing the growth, photosynthetic and calcification rates. In addition, the interaction of ocean acidification and decreased nitrate supply (projected for year 2100) had more negative synergistic effects on E. huxleyi physiology than all other two-way factorial manipulations, suggesting a linkage between the single dominant driver (nitrate) effects and interactive effects with other drivers. Simultaneous manipulation of all five environmental drivers to the projected year 2100 conditions had the largest negative effects on most of the physiological metrics. Furthermore, functional genes associated with inorganic carbon acquisition (RubisCO, AEL1 and delta CA) and calcification (CAX3, AEL1, PATP and NhaA2) were most down-regulated by the multiple driver manipulation, revealing linkages between responses of functional gene expression and associated physiological metrics.

In order to allow full comparability with other ocean acidification data sets, the R package seacarb (Gattuso et al, 2019) was used to compute a complete and consistent set of carbonate system variables, as described by Nisumaa et al. (2010). In this dataset the original values were archived in addition with the recalculated parameters (see related PI). The date of carbonate chemistry calculation by seacarb is 2020-07-07.