Dur3 and nrt2 genes in the bloom-forming dinoflagellate Prorocentrum minimum: Transcriptional responses to available nitrogen sources.

The increasing inflow of nitrogen (N) substrates into marine nearshore ecosystems induces proliferation of harmful algal blooms (HABs) of dinoflagellates, such as potentially toxic invasive species Prorocentrum minimum. In this study, we estimated the influence of NO3-, NH4+ and urea on transcription levels and urea transporter dur3 and nitrate transporter nrt2 genes expression in these dinoflagellates. We identified dur3 and nrt2 genes sequences in unannotated transcriptomes of P. minimum and other dinoflagellates presented in MMETSP database. Phylogenetic analysis showed that these genes of dinoflagellates clustered to the distinct clade demonstrating evolutionary relationship with the other known dur3 and nrt2 genes of microalgae. The evaluation of expression levels of dur3 and nrt2 genes by RT-qPCR revealed their sensitivity to input of the studied N sources. Dur3 expression levels were downregulated after the supplementation of additional N sources and were 1.7-2.6-fold lower than in the nitrate-grown culture. Nrt2 expression levels decreased 1.9-fold in the presence of NH4+. We estimated total RNA and DNA synthesis rates by the analysis of incorporation of 3H-thymidine and 3H-uridine in batch and continuous cultures. Addition of N compounds did not affect the DNA synthesis rates. Transcription levels increased up to 12.5-fold after the N supplementation in urea-limited treatments. Investigation of various nitrogen sources as biomarkers of dinoflagellate proliferation due to their differentiated impact on expression of dur3 and nrt2 genes and transcription rates in P. minimum cells allowed concluding about high potential of the studied parameters for future modeling of HABs under global N pollution.

MECHANICAL IMPACT ON THE CELL COVERING FINE STRUCTURE OF DINOFLAGELLATES PROROCENTRUM MINIMUM.

Complex cell coverings (amphiesma) of potentially toxic dinoflagellates Prorocentrum minimum include plasma membrane and flattened amphiesmal vesicles with thecal cellulose plates. Two largest thecal plates surround the major portion of dinoflagellate cell as shell valves. We have revealed that P. minimum cells appear to be extremely sensitive to the physical stress: even low speed centrifugation (1200 and 2000 g) leads to a dropping of old coverings shedding (ecdysis) and the formation of viable spheroplasts. Spheroplasts are surrounded only by the plasma membrane beneath which the new amphiesma is formed. These spheroplasts can be a convenient model system for investigation of numerous aspects of cell and molecular biology of the dinoflagellates.

[Mixotrophy in microorganisms: ecological and cytophysiological aspects].

Mixotrophy is the ability to combine autotrophic and heterotrophic modes of nutrition. It is widely spread in various microorganisms, particularly in such important plankton groups as dinoflagellates and cyanobacteria. Mixotrophy has a significant impact on our comprehension of the matter and energy flows in marine ecosystems, and therefore, it is an object of much attention for several recent decades. Nevertheless, the precise data on the balance of auto- and heterotrophy during the mixotrophic growth have been absent so far, which is due, first of all, to insufficient understanding of physiological and molecular ground of this phenomenon. In this review we discuss some ecological and cytophysiological aspects of investigation of mixotrophy in microorganisms as well as possible reasons for relatively slow progress in this area.