Transcriptomic Profiling of Genes Related to Cell Division, Metabolism and Toxin Production in the Dinoflagellate “Alexandrium fundyense”

Environmental conditions conducive to the explosive growth and formation of harmful algal blooms generally range from unique ability to efficiently utilize ambient nutrients, acquire light energy, and to defer grazing. In this dissertation, genetic potential for cell division, metabolism, and saxitoxin production in dinoflagellate “Alexandrium fundyense” (Atama Group I /Clade I) were studied by transcriptome profiling using 454 pyrosequencing. Transcriptome data were generated from a cultured strain (CCMP1719) and a natural bloom (in Long Island Sound) of “A. fundyense” collected over a 24 h period. More than 7.7 and 8.2 million dinoflagellate spliced leader-based 5’ expressed sequence tags were obtained for the laboratory culture and natural bloom respectively from four time points in the diel cycle representing G1, S, G2M of cell cycle phases and light/dark transition. Assembly of the reads yielded 87,273 unique genes for the natural bloom samples and 31,451 for the culture samples with ca. 25% of each assigned putative function. ESTs encoding majority of the enzymes involved in carbohydrate metabolism, oxidative stress response and toxin production were identified in “A. fundyense”. A total of 620 unique genes that encode putative membrane transporters were also identified. Transcript abundance quantified by reverse transcriptase quantitative PCR revealed the significant up-regulation of genes related to carbon fixation, saxitoxin production, nitrogen metabolism in the natural bloom with respect to those in the laboratory culture, indicating the nitrogen-depleted condition during the peak of the bloom and the versatility of “A. fundyense” in exploiting all possible sources of nitrogen to maintain the bloom. To further study the cell cycle regulation in this species, the full-length cDNA of a cyclin gene was isolated from “A. fundyense”, named Afcyc. The deduced protein sequence is closest to, and phylogenetically clusters with documented mitotic cyclin from dinoflagellate Lingulodinium polyedrum among cyclin B sequences from various eukaryotes. Afcyc transcript abundance was over 6-fold higher in the G2M phase than in other cell cycle phases, and showed a positive correlation with the percentage of cells in G2M phase, indicative of a mitotic cyclin, which could be a potential growth marker for monitoring the development of “A. fundyense” toxic blooms