Current estimates indicate that approximately half of the annual photosynthetic production of organic matter on Earth takes place in the euphotic zone of the water column in marine environments, making the ocean a major component of the global carbon cycle. Nearly all of the bloom-forming phytoplankton in the modern ocean, including the diatoms, are members of the chlorophyll c (Chl c) lineage. It is now well accepted that chlorophyll c-containing algae share a common ancestry where a eukaryote enveloped and domesticated a red algae, incorporating biochemical entities from both organisms. The subsequent diversification of this lineage has yielded an astounding diversity of ecologically dominant phytoplankton. Ultimately carbon export in marine ecosystems is governed and balanced by assimilation of nitrogen by phytoplankton. Among marine phytoplankton, diatoms are often the most responsive to mixing events where nutrient laden water is injected into the sunlit euphotic zone, yet the cellular basis for this competitive advantage is poorly understood.
The global state-of-knowledge concerning nitrogen metabolism in photosynthetic eukaryotes is largely based on biochemistry and genome sequence data from a few well-studied model green algae and vascular plants. Genome sequences, physiological and biochemical studies, and gene and protein expression data indicate that diatoms and other Chl c algae utilize a variety of biochemical components previously only observed in metazoans or bacteria. A complete metazoan-like urea cycle, for example, appears to function in concert with genes of bacterial and red or green algal origin in a functionally expanded and heretofore unobserved fashion. In metazoans the urea cycle has a function related to cellular detoxification and export of fixed nitrogen. The presence of the urea degrading enzyme urease strongly suggests an alternative function in diatoms.
This research is addressing the hypothesis that in marine diatoms, which are frequently subjected to nitrogen limitation, the urea cycle and associated mitochondrial enzymes function as an inorganic carbon and nitrogen recycling and repackaging hub; ultimately serving to redistribute catabolically derived NH4+ to arginine and other non-amino-acid nitrogen compounds. Based on recent advancements in diatom reverse genetics and other recently developed resources for genome enabled research, basic hypothesis concerning the role of mitochondria in nitrogen assimilation and metabolism in marine diatoms is being evaluated.
Broader Impacts Anthropogenic coastal loading of organic and inorganic nitrogen is expected to increase and climate change-induced shifts in oceanic mixed layer depth and mixing frequency are predicted to alter the delivery of nutrients into the euphotic zone. As a result, a key research focus is to facilitate prediction of how diatoms and other Chl c algae will respond to changing frequency and intensity of nutrient delivery events. This research will contribute to a more accurate depiction of nitrogen metabolism in marine algae, which is critically important for predictive ecosystem modeling.
As part of the research project, a high school biology teacher from the Escondido Union High School District in San Diego will be recruited to work on a specific topic related to the proposed research. Upon completion of his/her paid internship, in collaboration with the PIs, the teacher will design a classroom activity and curriculum installment related to the rapidly emerging field of marine genomics, also including associated topics in marine biogeochemistry, for use the following school year.
Current estimates indicate that approximately half of the annual photosynthetic production of organic matter on Earth takes place in the photic zone of the water column in marine environments; making the ocean a major component of the global carbon cycle. The prominence of diatoms in the contemporary ocean is such that the oxygen in every fifth breath take can be traced back to them. Further more, diatoms are a key component of the biological carbon pump that exports carbon to the ocean interior, contributing significantly to the long-term sequestration of atmospheric CO2. In addition to ecological considerations, diatoms occupy a supremely interesting and poorly understood position within the tree of life. It is now well accepted that chlorophyll c-containing algae share a common ancestry due to an ancient evolutionary event that resulted in the domestication of 'endosymbiotic' red algal cells, by an 'exosymbiotic' host, into what are now called secondary plastids. The subsequent diversification of this lineage has yielded an astounding diversity of ecologically dominant phytoplankton that manage pivotal roles in major geochemical processes (e.g., coastal upwelling, new production, and particle flux). Research completed through this project examined the evolutionary origins and biochemical function of genes and proteins that govern nitrogen nutrition and metabolic pathways in diatom cells. It appears that diatoms genomes are comprised of unique recombinations of typically segregated proteins (e.g., between plants, animals, and bacteria for example) and that this likely contributes to the ecological success of diatoms in the modern ocean. For example,diatom genomes encode a complete metazoan-like urea cycle, which appears to have been acquired from the exosymbiotic hosts that incorporates genes of bacterial and endosymbiont origin in a functionally expanded and heretofore unobserved fashion. Linkages between the urea cycle and other nitrogen processing proteins like glutamine synthase (GS III), arginine decarboxylase, and agmatinase have major metabolic and ecological implications for the basic nitrogen physiology of the diatom. Results from this project indicate that, in diatoms, the urea cycle has an anabolic function in contrast to animals where it has a more catabolic function. Specifically, in metazoans the urea cycle has a function related to cellular detoxification and export of fixed nitrogen. In marine diatoms, however, which are frequently subjected to nitrogen limitation, our data suggest that the urea cycle and associated enzymes function as an inorganic C and N recycling and repackaging hub; ultimately serving to redistribute catabolically derived NH4+ to arginine and other non-amino-acid nitrogen compounds. In concert, a complete glutamine synthase-glutamate synthase (GS-GOGAT) cycle in the diatom mitochondria allows for a rapid redistribution of nitrogen to amino-acids following the cessation of nutrient limitation. This research developed new methods for DNA delivery into diatoms and genetic manipulation of diatom genes and biochemistry. These techniques were used to examine the metabolic role of diatom mitochondria in overall cellular nitrogen metabolism. The diatom mitochondria 'houses' a variety of metabolic pathways, comprised of components that typically (in plants, animals, and bacteria) do not occur in the same organism. Together they facilitate numerous nitrogen transformations that permit rapid transitions between stationary phase (associated with nutrient exhaustion and minimal growth) and explosive exponential growth and nutrient consumption. The completed project fostered integration of important modern themes in genome biology, cellular metabolism, functional genomics and biogeochemistry, and provided novel insights into factors that control the distribution and nutrient biogeochemistry of diatoms in ocean systems. By extending development of functional genomics tools and resources for a model marine diatom, we significantly advanced the research potential of diatoms for applied biotechnology. As part of the completed research, we established a relationship with high school biology teachers at Orange Glen High School in the Escondido Union High School District in San Diego. Approximately 74% of the student body is Hispanic and 65% of the students are enrolled in the free/reduced price lunch program. We recruited two high school teachers from the Escondido Union District to participate in this research project. A presentation on their research was delivered at the National Association of Biology Teachers annual meeting.
