Nitrate Reductase Knockout Uncouples Nitrate Transport from Nitrate Assimilation and Drives Repartitioning of Carbon Flux in a Model Pennate Diatom

The ecological prominence of diatoms in the ocean environment largely results from their superior competitive ability for dissolved nitrate (NO 3 − ). To investigate the cellular and genetic basis of diatom NO 3 − assimilation, we generated a knockout in the nitrate reductase gene ( NR -KO) of the model pennate diatom Phaeodactylum tricornutum . In NR -KO cells, N-assimilation was abolished although NO 3 − transport remained intact. Unassimilated NO 3 − accumulated in NR -KO cells, resulting in swelling and associated changes in biochemical composition and physiology. Elevated expression of genes encoding putative vacuolar NO 3 − chloride channel transporters plus electron micrographs indicating enlarged vacuoles suggested vacuolar storage of NO 3 − . Triacylglycerol concentrations in the NR -KO cells increased immediately following the addition of NO 3 − , and these increases coincided with elevated gene expression of key triacylglycerol biosynthesis components. Simultaneously, induction of transcripts encoding proteins involved in thylakoid membrane lipid recycling suggested more abrupt repartitioning of carbon resources in NR -KO cells compared with the wild type. Conversely, ribosomal structure and photosystem genes were immediately deactivated in NR -KO cells following NO 3 − addition, followed within hours by deactivation of genes encoding enzymes for chlorophyll biosynthesis and carbon fixation and metabolism. N-assimilation pathway genes respond uniquely, apparently induced simultaneously by both NO 3 − replete and deplete conditions.