Transcriptional Responses of Karenia brevis to Phosphorus Depletion and Addition

The role of coastal nutrient sources in the persistence of Karenia brevis red tides in coastal waters of Florida is currently a contentious issue that warrants investigation into the nutrient physiology of this dinoflagellate species. The molecular mechanisms by which dinoflagellates respond to nutrient availability are essentially unexplored. The current study sought to determine if the transcriptome of K. brevis is responsive to phosphorus and informative of nutrient status. Stationary phase P-limited cultures did not exhibit up-regulation of transcripts considered to be hallmark of P depletion relative to nutrient-replete stationary phase cultures, despite rapid growth upon addition of the limiting nutrient. To study transcriptome responses to nutrient addition, the limiting nutrient was added to depleted cells and changes in global gene expression were assessed over the first 48 hours following nutrient addition. P-addition resulted in significant changes in approximately 4% of genes on the microarray, using a significance cutoff of 1.7-fold and p<10-4. By far, the earliest responding genes were dominated by pentatricopeptide repeat (PPR) proteins, which increased in expression up to 3-fold by 1 h following nutrient addition. PPR proteins are nuclear encoded proteins involved in chloroplast and mitochondria RNA processing. Correspondingly, other functions enriched in response to both nutrients were photosystem and ribosomal genes. Transcriptomic responses to the addition of P reflected primarily chloroplast functions. Even the earliest responding transcripts, those encoding PPR proteins, possess a 5’ trans-spliced leader sequence, suggesting that the observed rapid response of the transcriptome may be achieved, in part, through post-transcriptional mechanisms. Cultures were acclimated to 0.1 µM phosphate for a minimum of two months during which time cultures were transferred weekly in log phase. For addition experiments, nutrient replete and P-limited 1L cultures were grown to stationary phase. (Day 12) Using sodium phosphate, 168 µM PO4 was added to stationary phase cultures. Nutrient replete cultures and P-limited cultures were harvested at the time of nutrient addition (Time 0; n=3) and total RNA extracted. Following phosphorus addition, triplicate cultures were harvested at 1, 4, 24, and 48 h post-addition and total RNA extracted. One color arrays were then run on all biological replicates (n=3 at each time point).