Visualizing in situ translational activity for identifying and sorting slow-growing archaeal−bacterial consortia
Abstract
To understand the biogeochemical roles of microorganisms in the environment, it is important to determine when and under which conditions they are metabolically active. Bioorthogonal noncanonical amino acid tagging (BONCAT) can reveal active cells by tracking the incorporation of synthetic amino acids into newly synthesized proteins. The phylogenetic identity of translationally active cells can be determined by combining BONCAT with rRNA-targeted fluorescence in situ hybridization (BONCAT-FISH). In theory, BONCAT-labeled cells could be isolated with fluorescence-activated cell sorting (BONCAT-FACS) for subsequent genetic analyses. Here, in the first application, to our knowledge, of BONCAT-FISH and BONCAT-FACS within an environmental context, we probe the translational activity of microbial consortia catalyzing the anaerobic oxidation of methane (AOM), a dominant sink of methane in the ocean. These consortia, which typically are composed of anaerobic methane-oxidizing archaea (ANME) and sulfate-reducing bacteria, have been difficult to study due to their slow in situ growth rates, and fundamental questions remain about their ecology and diversity of interactions occurring between ANME and associated partners. Our activity-correlated analyses of > 16,400 microbial aggregates provide the first evidence, to our knowledge, that AOM consortia affiliated with all five major ANME clades are concurrently active under controlled conditions. Surprisingly, sorting of individualmore »
- Authors:
- Publication Date:
- Research Org.:
- Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER); National Science Foundation (NSF); Austrian Science Fund (FWF); Gordon and Betty Moore Foundation
- OSTI Identifier:
- 1259913
- Alternate Identifier(s):
- OSTI ID: 1379511
- Grant/Contract Number:
- AC02-05CH11231; OCE-0825791; GBMF3780
- Resource Type:
- Published Article
- Journal Name:
- Proceedings of the National Academy of Sciences of the United States of America
- Additional Journal Information:
- Journal Name: Proceedings of the National Academy of Sciences of the United States of America Journal Volume: 113 Journal Issue: 28; Journal ID: ISSN 0027-8424
- Publisher:
- National Academy of Sciences, Washington, DC (United States)
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; activity-based cell-sorting; BONCAT; click chemistry; ecophysiology; single-cell microbiology
Citation Formats
Hatzenpichler, Roland, Connon, Stephanie A., Goudeau, Danielle, Malmstrom, Rex R., Woyke, Tanja, and Orphan, Victoria J. Visualizing in situ translational activity for identifying and sorting slow-growing archaeal−bacterial consortia. United States: N. p., 2016.
Web. doi:10.1073/pnas.1603757113.
Hatzenpichler, Roland, Connon, Stephanie A., Goudeau, Danielle, Malmstrom, Rex R., Woyke, Tanja, & Orphan, Victoria J. Visualizing in situ translational activity for identifying and sorting slow-growing archaeal−bacterial consortia. United States. https://doi.org/10.1073/pnas.1603757113
Hatzenpichler, Roland, Connon, Stephanie A., Goudeau, Danielle, Malmstrom, Rex R., Woyke, Tanja, and Orphan, Victoria J. Tue .
"Visualizing in situ translational activity for identifying and sorting slow-growing archaeal−bacterial consortia". United States. https://doi.org/10.1073/pnas.1603757113.
@article{osti_1259913,
title = {Visualizing in situ translational activity for identifying and sorting slow-growing archaeal−bacterial consortia},
author = {Hatzenpichler, Roland and Connon, Stephanie A. and Goudeau, Danielle and Malmstrom, Rex R. and Woyke, Tanja and Orphan, Victoria J.},
abstractNote = {To understand the biogeochemical roles of microorganisms in the environment, it is important to determine when and under which conditions they are metabolically active. Bioorthogonal noncanonical amino acid tagging (BONCAT) can reveal active cells by tracking the incorporation of synthetic amino acids into newly synthesized proteins. The phylogenetic identity of translationally active cells can be determined by combining BONCAT with rRNA-targeted fluorescence in situ hybridization (BONCAT-FISH). In theory, BONCAT-labeled cells could be isolated with fluorescence-activated cell sorting (BONCAT-FACS) for subsequent genetic analyses. Here, in the first application, to our knowledge, of BONCAT-FISH and BONCAT-FACS within an environmental context, we probe the translational activity of microbial consortia catalyzing the anaerobic oxidation of methane (AOM), a dominant sink of methane in the ocean. These consortia, which typically are composed of anaerobic methane-oxidizing archaea (ANME) and sulfate-reducing bacteria, have been difficult to study due to their slow in situ growth rates, and fundamental questions remain about their ecology and diversity of interactions occurring between ANME and associated partners. Our activity-correlated analyses of > 16,400 microbial aggregates provide the first evidence, to our knowledge, that AOM consortia affiliated with all five major ANME clades are concurrently active under controlled conditions. Surprisingly, sorting of individual BONCAT-labeled consortia followed by whole-genome amplification and 16S rRNA gene sequencing revealed previously unrecognized interactions of ANME with members of the poorly understood phylum Verrucomicrobia. This finding, together with our observation that ANME-associated Verrucomicrobia are found in a variety of geographically distinct methane seep environments, suggests a broader range of symbiotic relationships within AOM consortia than previously thought.},
doi = {10.1073/pnas.1603757113},
journal = {Proceedings of the National Academy of Sciences of the United States of America},
number = 28,
volume = 113,
place = {United States},
year = {Tue Jun 28 00:00:00 EDT 2016},
month = {Tue Jun 28 00:00:00 EDT 2016}
}
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