FY2010 - Oak Ridge National Laboratory

Director’s R&D Fund—
Systems Biology and the Environment
interaction will be dissected using comparative genomics, gene expression, and proteomics with
Ignicoccus species that are not symbiotic with N. equitans. This will allow us to identify candidate genes
that mediate cell–cell communication and to study physical interaction processes at the membrane level.
Structural and molecular dynamics characteristics of selected proteins and protein complexes will be
investigated using computational and experimental methods to understand the mechanisms of intercellular
interaction. General principles will be derived on interspecific relationships and how they are mediated at
the genomic level.
Mission Relevance
Few if any microbes in the environment live in isolation of each other. Microbial metabolic synergies and
specialized syntrophic relationships are responsible for numerous environmental processes of significant
importance for the DOE mission including anaerobic methane oxidation, biotransformation of xenobiotics
and heavy metals. Holistic studies of microbial interspecies relationships are still lacking due to
difficulties in maintaining symbiotic and syntrophic systems in the laboratory. Our research is aimed at
deciphering physiological, molecular, and genomic mechanisms of interaction between two species. By
combining an interdisciplinary, systems biology approach to study this system, we will establish a
platform to study more complex syntrophic associations and will derive fundamental principles on how
microbes interact. This research is of high importance to the DOE Office of Biological and Environmental
Science (BER) Genomic Science Program (formerly Genomics: _GTL) focus area and will be part of
future ORNL Scientific Focus Areac research or collaborative university-led Genomic Science
solicitations. Integrated systems biology research on syntrophic/symbiotic relationships is also of high
relevance to microbial communities of interest to the National Science Foundation (NSF) programs,
National Aeronautics and Space Administration (NASA)-–Astrobiology, and the National Institutes of
Health (NIH) Microbiome program.
Results and Accomplishments
Objective 1. Genome sequencing of Ignicoccus islandicus and I. pacificus. We have obtained draft
sequences for both genomes using a combination of 454 Titanium 8kb paired-insert library and Illumina
paired end library sequencing. Analysis is ongoing for genome closure.
Detection and isolation of novel nanoarchaea. (1) Detection of nanoarchaea in environmental samples,
using universal and specific primers. A large number of samples (>60) were analyzed using 454
sequencing and specific full length sequencing of nanoarchaea 16S rDNA, including deep sea
hydrothermal samples from the Mid Atlantic Ridge, Guaymas, Lao, Juan de Fuca, Indian Ocean,
numerous samples from terrestrial sites around the world. A manuscript detailing the results of these
studies is in preparation. (2) Enrichments for cultivation. We have set up batch cultures and continuous
fermentation systems to maintain, enrich and isolate novel nanoarchea from both marine and terrestrial
sites. (3) We have developed an affinity isolation method to be used for isolation of novel nanoarchaea.
The system uses antibody against N. equitans that is coupled to paramagnetic beads. This system has been
tested on I. hospitalis-N. equitans and is being applied to isolate novel nanoarchea species from
environmental samples or from enrichments. The same approach could be used for the isolation of other
organisms as well.
Objective 2. Functional genomics of the Ignicoccus–Nanoarchaeum interaction. We have completed
in-depth proteomic and microarray studies of cultures at various stages in the association. The data is
being analyzed and two publications are planned, one is in advanced manuscript stage. We are in the
process of developing an RNA sequencing approach to analyze general transcripts as well as small
regulatory RNAs.
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