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A mutant (‘lab strain’) of the hyperthermophilic archaeon Pyrococcus furiosus, lacking flagella, has unusual growth physiology

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Abstract

A mutant (‘lab strain’) of the hyperthermophilic archaeon Pyrococcus furiosus DSM3638 exhibited an extended exponential phase and atypical cell aggregation behavior. Genomic DNA from the mutant culture was sequenced and compared to wild-type (WT) DSM3638, revealing 145 genes with one or more insertions, deletions, or substitutions (12 silent, 33 amino acid substitutions, and 100 frame shifts). Approximately, half of the mutated genes were transposases or hypothetical proteins. The WT transcriptome revealed numerous changes in amino acid and pyrimidine biosynthesis pathways coincidental with growth phase transitions, unlike the mutant whose transcriptome reflected the observed prolonged exponential phase. Targeted gene deletions, based on frame-shifted ORFs in the mutant genome, in a genetically tractable strain of P. furiosus (COM1) could not generate the extended exponential phase behavior observed for the mutant. For example, a putative radical SAM family protein (PF2064) was the most highly up-regulated ORF (>25-fold) in the WT between exponential and stationary phase, although this ORF was unresponsive in the mutant; deletion of this gene in P. furiosus COM1 resulted in no apparent phenotype. On the other hand, frame-shifting mutations in the mutant genome negatively impacted transcription of a flagellar biosynthesis operon (PF0329-PF0338).Consequently, cells in the mutant culture lacked flagella and, unlike the WT, showed minimal evidence of exopolysaccharide-based cell aggregation in post-exponential phase. Electron microscopy of PF0331-PF0337 deletions in P. furiosus COM1 showed that absence of flagella impacted normal cell aggregation behavior and, furthermore, indicated that flagella play a key role, beyond motility, in the growth physiology of P. furiosus.

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Acknowledgments

This work was supported by a grant from National Science Foundation (CBT-0730091) to RMK. Construction and characterization of the deletion mutants was funded by a grant to MWA (FG05-95ER20175) from the Chemical Sciences, Geosciences and Biosciences Division of the Office of Basic Energy Sciences, Office of Science, and U.S. Department of Energy. The authors are grateful to Dr. Jennifer Schaff, Genome Sciences Laboratory at NCSU, for her help with genome sequencing and Dr. Michael J. Dykstra, LAELOM, College of Veterinary Medicine, NCSU, for help with TEM and SEM imaging.

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  1. Derrick L. Lewis

    Present address: Novozymes Biologicals, Salem, VA, US

Authors and Affiliations

  1. Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, EB-1,911 Partners Way, Raleigh, NC, 27695-7905, US

    Derrick L. Lewis, Jaspreet S. Notey, Andrew J. Loder & Robert M. Kelly

  2. Department of Biochemistry and Molecular Biology, University of Georgia, Athens, GA, 30602, US

    Sanjeev K. Chandrayan, Gina L. Lipscomb & Michael W. W. Adams

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  1. Derrick L. Lewis
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Correspondence to Robert M. Kelly.

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Communicated by H. Atomi.

D. L. Lewis, J. S. Notey contributed equally to this work.

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Lewis, D.L., Notey, J.S., Chandrayan, S.K. et al. A mutant (‘lab strain’) of the hyperthermophilic archaeon Pyrococcus furiosus, lacking flagella, has unusual growth physiology. Extremophiles 19, 269–281 (2015). https://doi.org/10.1007/s00792-014-0712-3

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