Targeted hypermutation of putative antigen sensors in multicellular bacteria
Edited by Marlene Belfort, University at Albany, State University of New York, Albany, NY; received September 29, 2023; accepted January 10, 2024
Significance
To defend themselves against pathogens, bacteria employ a wide range of conflict systems, some of which are enriched in multicellular bacteria. Here, we show that numerous multicellular bacteria use related diversity-generating retroelements (DGRs) to diversify such putative conflict systems. Error-prone reverse transcription in DGRs introduces random, targeted mutations and rapid diversification. We used Thiohalocapsa PB-PSB1, a member of multicellular bacterial consortia, to study this association between conflict systems and DGRs. We characterized the natural diversity of PB-PSB1 DGRs and propose they function as hypervariable antigen sensors. If their role in pathogen defense is confirmed, accumulation of these DGR-diversified systems in multicellular bacteria would suggest that rapidly diversifying immune systems confer important fitness advantages for the evolution of multicellularity.
Abstract
Diversity-generating retroelements (DGRs) are used by bacteria, archaea, and viruses as a targeted mutagenesis tool. Through error-prone reverse transcription, DGRs introduce random mutations at specific genomic loci, enabling rapid evolution of these targeted genes. However, the function and benefits of DGR-diversified proteins in cellular hosts remain elusive. We find that 82% of DGRs from one of the major monophyletic lineages of DGR reverse transcriptases are encoded by multicellular bacteria, which often have two or more DGR loci in their genomes. Using the multicellular purple sulfur bacterium Thiohalocapsa sp. PB-PSB1 as an example, we characterized nine distinct DGR loci capable of generating 10282 different combinations of target proteins. With environmental metagenomes from individual Thiohalocapsa aggregates, we show that most of PB-PSB1’s DGR target genes are diversified across its biogeographic range, with spatial heterogeneity in the diversity of each locus. In Thiohalocapsa PB-PSB1 and other bacteria hosting this lineage of cellular DGRs, the diversified target genes are associated with NACHT-domain anti-phage defenses and putative ternary conflict systems previously shown to be enriched in multicellular bacteria. We propose that these DGR-diversified targets act as antigen sensors that confer a form of adaptive immunity to their multicellular consortia, though this remains to be experimentally tested. These findings could have implications for understanding the evolution of multicellularity, as the NACHT-domain anti-phage systems and ternary systems share both domain homology and conceptual similarities with the innate immune and programmed cell death pathways of plants and metazoans.
Data, Materials, and Software Availability
All new sequencing data are available in NCBI GenBank under BioProject PRJNA1019683 (93) and JGI’s IMG database (3300056627, 3300056928, 3300056818) (94–96). Accession numbers are provided in Dataset S5. Code has been deposited in GitHub (https://doi.org/10.5281/zenodo.10569842) (76).
Acknowledgments
We thank L. Aravind for providing sequence alignments and hmm profiles for putative conflict systems. We also thank José T. Saavedra and Scott Chimileski for the photographs in Fig. 1. Some figure panels were created with BioRender.com. This work was supported by a Whitman Fellowship to E.G.W. from the Marine Biological Laboratory. Use was made of computational facilities purchased with NSF funds (CNS-1725797) and administered by the Center for Scientific Computing, which is supported by the California NanoSystems Institute and the Materials Research Science and Engineering Center (MRSEC; NSF DMR 2308708) at UCSB. Research was sponsored by the U.S. Army Research Office and accomplished under cooperative agreement W911NF-19-2-0026 and W911NF-19-D-0001 for the Institute for Collaborative Biotechnologies. Long read sequencing was supported by a JGI New Investigator award to E.G.W. (508543). An open-access license has been selected for this work.
Author contributions
H.D., A.R.E., and E.G.W. designed research; H.D., A.R.E., E.N.J., G.E.L., O.X.C., D.L.V., M.A.O., and E.G.W. performed research; H.D., A.G., B.G.P., and E.G.W. contributed new analytic tools; H.D., A.R.E., B.G.P., and E.G.W. analyzed data; and H.D., A.R.E., and E.G.W. wrote the paper.
Competing interests
The authors declare no competing interest.
Supporting Information
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Copyright © 2024 the Author(s). Published by PNAS. This open access article is distributed under Creative Commons Attribution License 4.0 (CC BY).
Data, Materials, and Software Availability
All new sequencing data are available in NCBI GenBank under BioProject PRJNA1019683 (93) and JGI’s IMG database (3300056627, 3300056928, 3300056818) (94–96). Accession numbers are provided in Dataset S5. Code has been deposited in GitHub (https://doi.org/10.5281/zenodo.10569842) (76).
Submission history
Received: September 29, 2023
Accepted: January 10, 2024
Published online: February 14, 2024
Published in issue: February 27, 2024
Change history
March 6, 2024: The author names and affiliations have been updated to correct a production error. The Acknowledgments have been updated to correct a typographical error. Previous version (February 14, 2024)
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Acknowledgments
We thank L. Aravind for providing sequence alignments and hmm profiles for putative conflict systems. We also thank José T. Saavedra and Scott Chimileski for the photographs in Fig. 1. Some figure panels were created with BioRender.com. This work was supported by a Whitman Fellowship to E.G.W. from the Marine Biological Laboratory. Use was made of computational facilities purchased with NSF funds (CNS-1725797) and administered by the Center for Scientific Computing, which is supported by the California NanoSystems Institute and the Materials Research Science and Engineering Center (MRSEC; NSF DMR 2308708) at UCSB. Research was sponsored by the U.S. Army Research Office and accomplished under cooperative agreement W911NF-19-2-0026 and W911NF-19-D-0001 for the Institute for Collaborative Biotechnologies. Long read sequencing was supported by a JGI New Investigator award to E.G.W. (508543). An open-access license has been selected for this work.
Author contributions
H.D., A.R.E., and E.G.W. designed research; H.D., A.R.E., E.N.J., G.E.L., O.X.C., D.L.V., M.A.O., and E.G.W. performed research; H.D., A.G., B.G.P., and E.G.W. contributed new analytic tools; H.D., A.R.E., B.G.P., and E.G.W. analyzed data; and H.D., A.R.E., and E.G.W. wrote the paper.
Competing interests
The authors declare no competing interest.
Notes
This article is a PNAS Direct Submission.
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Targeted hypermutation of putative antigen sensors in multicellular bacteria, Proc. Natl. Acad. Sci. U.S.A.
121 (9) e2316469121,
https://doi.org/10.1073/pnas.2316469121
(2024).
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