Kleiner, F. H. | Search bioRxiv

bioRxiv2022

Establishment of a novel set of vectors for transformation of the dinoflagellate Amphidinium carterae

Peridinin-containing dinoflagellate algae have a chloroplast genome formed from plasmid-like minicircles. This fragmented genome has allowed us to develop a genetic modification methodology involving the use of biolistics to introduce artificial minicircles in Amphidinium carterae (Nimmo et al., 2019). The previously reported artificial minicircles were based on native minicircles containing either the psbA or atpB gene. Each artificial minicircle allowed expression of a single selectable marker instead of psbA or atpB. Here, we present two further artificial minicircles for use in transformation of A. carterae. One is based on the petD minicircle, allowing the expression of a single selectable marker. The second is based on the two-gene minicircle originally containing atpA and petB, and allows the dual expression of a selectable marker and a gene of interest. Our research suggest that all of the 20 or so minicircles in A. carterae are suitable for adaptation as artificial minicircles, allowing for the simultaneous introduction of multiple genes.

plant biology↗

bioRxiv2022

Cold-induced cyt elevations function to support osmoregulation in marine diatoms

Diatoms are a group of microalgae that are important primary producers in a range of open ocean, freshwater and intertidal environments. The latter can experience significant long- and short-term variability in temperature, from seasonal variations to rapid temperature shifts caused by tidal immersion and emersion. As temperature is a major determinant in the distribution of diatom species, their temperature sensory and response mechanisms likely have important roles in their ecological success. We have examined the mechanisms diatoms use to sense rapid changes in temperature, such as those experienced in the intertidal zone. We find that the diatoms Phaeodactylum tricornutum and Thalassiosira pseudonana exhibit a transient cytosolic Ca2+ ([Ca2+]cyt) elevation in response to rapid cooling, similar to those observed in plant and animal cells. However, [Ca2+]cyt elevations were not observed in response to rapid warming. The kinetics and magnitude of cold-induced [Ca2+]cyt elevations correlate with the rate of temperature decrease. We do not find a role for the [Ca2+]cyt elevations in enhancing cold tolerance, but show that cold shock induces a Ca2+-dependent K+ efflux and reduces mortality of P. tricornutum during a simultaneous hypo-osmotic shock. As inter-tidal diatom species may routinely encounter simultaneous cold and hypo-osmotic shocks during tidal cycles, we propose that cold-induced Ca2+ signalling interacts with osmotic signalling pathways to aid in the regulation of cell volume. Our findings provide insight into the nature of temperature perception in diatoms and highlight that cross-talk between signalling pathways may play an important role in their cellular responses to multiple simultaneous stressors.

plant biology↗