2013 Annual Science Report
Arizona State University Reporting | SEP 2012 – AUG 2013
Stoichiometry of Life - Task 1 - Laboratory Studies in Biological Stoichiometry
This project component involves a set of studies of microorganisms with which we are trying to better understand how living things use chemical elements (nitrogen, phosphorus, iron, etc.) and how they cope, in a physiological sense, with shortages of such elements. For example, how does the “elemental recipe of life” change when an organism is starved for phosphorus or nitrogen or iron? Is this change similar for different species of microorganisms? Are the changes the same if the organism is limited by a different key nutrient? Furthermore, how does an organism shift its patterns of gene expression when it is starved by various nutrients? This will help in interpreting studies of gene expression in natural environments.
Effects of nutrient limitation on microbial stoichiometry and metabolism:
Batch cultures of the cyanobacterium, Synechocystis sp. PCC 6803 were grown under different nutrient conditions: Complete (media rich in all nutrients; N/P ratio of 100), Nitrogen-limited; media with low concentration of N; N/P ratio of 10), Phosphorus-limited; (media with low concentrations of P; N/P ratio of 1000); and Iron-limited (media with low concentration of Fe; 10 times less Fe than complete media). During the exponential phase of growth, all cultures were subjected to analysis of elemental stoichiometry (C:N:P:S:metals), biological macromolecule composition (DNA, RNA, protein, chlorophyll), transcriptome profiles, and metabolome profiles. The elemental composition of cultures was determined via EA-IRMS (C and N) and ICP-MS (P and metals) and biological macromolecule composition using fluorescent or spectrophotometric methods. Metabolome profiling was conducted using LC-MS Q-TOF (Liquid Chromatography with Time-of-Flight Mass Spectrometry) and NMR spectroscopy analyses. Transcriptome profiling was conducted by Ion Torrent sequencing of cDNA derived from rRNA-subtracted RNA extractions. These analyses show that phosphorus and iron-limited cultures significantly shifted their elemental and biological macromolecule composition. Cultures under both nutrient limitations had (1) increased C:N, C:P, and N:P ratios, (2) decreased RNA:DNA and RNA:Protein ratios, and (3) iron-limited cultures had lower Fe:C ratios. Metal:C ratios showed an unexpected trend with phosphorus-limitation with significant increases in this ratio for some trace metals. Changes in biomass composition were also reflected in metabolomic and transcriptomic profiles, with the metabolome showing changes in the concentrations of organic acid metabolites and carbohydrates under nutrient limitation. Transcriptomic analyses also revealed a number of significantly differentially expressed genes encoding hypothetical proteins that may be targets for future experimental manipulation to determine their roles in responses to nutrient limitation. Results from elemental, biological macromolecule, metabolomic, and transcriptomic analyses are currently being integrated. Manuscripts describing the results of these experiments are in preparation.
Interactions of molybdenum with other biogeochemical cycles:
In order to determine the regulation of the putative Mo storage protein “Mop” in freshwater heterocystous cyanobacteria, Glass and undergraduate research assistant Eric Hughes performed various experiments manipulating Mo supplies with the model organism Nostoc sp. PCC 7120. A manuscript reporting results from this study has been published in Advances in Microbiology.
PROJECT INVESTIGATORS:James Elser
PROJECT MEMBERS:Ariel Anbar
RELATED OBJECTIVES:Objective 5.2
Co-evolution of microbial communities
Biochemical adaptation to extreme environments
Effects of environmental changes on microbial ecosystems
Adaptation and evolution of life beyond Earth