A New Interdisciplinary Frontier: Merging Molecular Biology with Isotopic Geochemistry

Harvard University, Cambridge MA

Investigators

Abstract

The recently appreciated scope of microbial diversity hints at vast hidden metabolic complexity in the environment. These processes govern the transfer of carbon and energy in natural systems. However, less than one percent of all known prokaryotes can be grown in pure culture. New, culture-independent methods are required to investigate the ecological roles and metabolic pathways of uncultivable prokaryotes. This project will demonstrate that carbon isotopic analysis of nucleic acids, specifically ribosomal RNA (rRNA), can be used to study microbial metabolism. The native rRNA in any sample is molecular biology's analogue to the "biomarker" concept of geochemistry. The key to realizing this important step is the recent development of a viable analytical method for RNA separation and isotopic analysis. Intact, species-specific rRNA is separated from mixtures of total RNA using magnetic bead capture methods. The pure material is transferred to a moving-wire interface attached to an isotope-ratio mass spectrometer. This represents a truly interdisciplinary achievement, as it permits the analysis of 13C/12C ratios of nanogram-sized samples of rRNA. It is possible to obtain both isotopic and phylogenetic information, simultaneously, from any natural sample. Scientific Goals This preliminary study will focus on topics of recent interest in geobiology. (i) What is the metabolic pathway of carbon assimilation in marine planktonic crenarchaeota? Is the hypothesis that this organism uses the 3-hydroxypropionate (3-HP) pathway correct? Is the magnitude of this isotopic fractionation the same in nature as it is in laboratory cultures? If not, what might be learned about the role of the 3-HP pathway in natural communities? (ii) Can we identify the major pathway to CH4 in sedimentary and geothermal environments? To what extent does anaerobic methane oxidation (AMO) contribute to total deep biosphere productivity? Is it possible to distinguish metabolic differences between different microbial taxa involved in AMO? The project fosters collaboration among research institutions and between biologists and geoscientists. In addition to the P.I., there are five individuals representing three research laboratories who also will participate in some aspect of the project. This work also supports the mission of NSF to foster educational development at the graduate and undergraduate levels. A graduate student Ph.D. thesis will result directly from this work; and some of the sampling will be performed by undergraduate students as the field component of a required academic course.