Single-cell microscopy reveals that levels of cyclic di-GMP vary among bacillus subtilis subpopulations

Cordelia A. Weiss, Jakob A. Hoberg, Kuanqing Liu, Benjamin P. Tu, Wade C. Winkler

Research output: Contribution to journalArticlepeer-review

21 Scopus citations


The synthesis of signaling molecules is one strategy bacteria employ to sense alterations in their environment and rapidly adjust to those changes. In Gramnegative bacteria, bis-(3=-5=)-cyclic dimeric GMP (c-di-GMP) regulates the transition from a unicellular motile state to a multicellular sessile state. However, c-di-GMP signaling has been less intensively studied in Gram-positive organisms. To that end, we constructed a fluorescent yfp reporter based on a c-di-GMP-responsive riboswitch to visualize the relative abundance of c-di-GMP for single cells of the Gram-positive model organism Bacillus subtilis. Coupled with cell-type-specific fluorescent reporters, this riboswitch reporter revealed that c-di-GMP levels are markedly different among B. subtilis cellular subpopulations. For example, cells that have made the decision to become matrix producers maintain higher intracellular c-di-GMP concentrations than motile cells. Similarly, we find that c-di-GMP levels differ between sporulating and competent cell types. These results suggest that biochemical measurements of c-di- GMP abundance are likely to be inaccurate for a bulk ensemble of B. subtilis cells, as such measurements will average c-di-GMP levels across the population. Moreover, the significant variation in c-di-GMP levels between cell types hints that c-di-GMP might play an important role during B. subtilis biofilm formation. This study therefore emphasizes the importance of using single-cell approaches for analyzing metabolic trends within ensemble bacterial populations.

Original languageEnglish (US)
Article numbere00247-19
JournalJournal of bacteriology
Issue number16
StatePublished - Aug 2019
Externally publishedYes


  • Bacillus subtilis
  • Biofilm
  • Cyclic di-GMP
  • Microscopy
  • Riboswitch

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology


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