Temperate viruses can become dormant in their host cells, a process called lysogeny. In every infection, such viruses decide between the lytic and the lysogenic cycles, that is, whether to replicate and lyse their host or to lysogenize and keep the host viable. Here we show that viruses (phages) of the SPbeta group use a small-molecule communication system to coordinate lysis–lysogeny decisions. During infection of its Bacillus host cell, the phage produces a six amino-acids-long communication peptide that is released into the medium. In subsequent infections, progeny phages measure the concentration of this peptide and lysogenize if the concentration is sufficiently high. We found that different phages encode different versions of the communication peptide, demonstrating a phage-specific peptide communication code for lysogeny decisions. We term this communication system the ‘arbitrium’ system, and further show that it is encoded by three phage genes: aimP, which produces the peptide; aimR, the intracellular peptide receptor; and aimX, a negative regulator of lysogeny. The arbitrium system enables a descendant phage to ‘communicate’ with its predecessors, that is, to estimate the amount of recent previous infections and hence decide whether to employ the lytic or lysogenic cycle.
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Extended data figures and tables
Extended Data Figures
- Extended Data Figure 1: Specificity of conditioned media. (241 KB)
Conditioned media were prepared using initial infection of B. subtilis 168 by four phages: phi3T, phi105, phi29 and rho14. Presented are growth curves of B. subtilis 168 in the different media, infected with each phage at MOI = 0.1: phi3T (a), phi105 (b), phi29 (c) and rho14 (d). Data represent average of 3 replicates, and error bars represent s.e.
- Extended Data Figure 2: Proteinase K treatment reduces the effect of conditioned medium. (188 KB)
Growth curves of B. subtilis 168 infected by phi3T at MOI = 0.1, in control and conditioned media, with and without pre-treatment with proteinase K. Data represent average of 3 technical replicates, and error bars represent s.e.
- Extended Data Figure 3: Mass spectrometry verifies the presence of the SAIRGA peptide in conditioned medium. (116 KB)
Presented are extracted ion chromatograms of targeted mass spectrometry analysis experiment. B2, Y3, Y4 and Y5 refer to fragmentation products of the peptides made by the instrument in the tandem mass spectrometry process, with the expected m/z ion masses indicated. The y axis represents the ion intensity of each fragment ion (arbitrary units). a, Reference synthesized SAIRGA peptide at 100nM concentration in LB. b, Control medium from B. subtilis 168. c, phi3T-infected B. subtilis 168. d, B. subtilis 168 expressing dCas9 with a spacer targeting aimP. e, phi3T-infected B. subtilis 168 expressing dCas9 with a spacer targeting aimP. Arrowhead depicts the expected retention time of the SAIRGA peptide (a, c).
- Extended Data Figure 4: 5 aa versions of the 6 aa arbitrium peptide do not guide lysogeny. (92 KB)
Growth curves of B. subtilis 168 infected by phi3T at MOI = 0.1, in LB media supplemented with synthesized SAIRGA, AIRGA or SAIRG peptide. Shown is average of 3 biological replicates, each with 3 technical replicates. Error bars represent s.e.
- Extended Data Figure 5: Exposure of AimR to SAIRGA peptide reduces propensity for dimerization. (141 KB)
Purified AimR was eluted from a gel-filtration column, dialysed, then mixed with SAIRGA peptide (final concentration, 100 μM) dissolved in water or with equal amount of volume without peptide and incubated at room temperature for 5 min. The protein samples were then mixed for 30 min with different concentrations of the crosslinker BS3 bis(sulfosuccinimidyl)suberate. Presented are electrophoresis results analysed using the TapeStation instrument (Agilent Technologies), showing that in the absence of peptide, purified AimR tends to preferentially be crosslinked into dimers, whereas in the presence of the SAIRGA peptide, dimerization is substantially reduced.
- Extended Data Figure 6: Phage gene expression 20 min after infection. (53 KB)
Each dot represents a single phage gene. Axes represent average RNA-seq read count per gene from 3 replicates, after normalization to control for RNA-seq library size. x axis, expression when phage infection was in the presence of 1 μM of SAIRGA peptide; y axis, no peptide.
- Extended Data Figure 7: Expression of the arbitrium locus in phage SPbeta during infection. (193 KB)
RNA-seq coverage of the arbitrium locus at 20 min after infection, with (black) or without (green) 1 μM of synthesized GMPRGA peptide in the medium. RNA-seq coverage was normalized to the number of reads mapped to the phage genome in each RNA-seq library. Shown are two replicates of the experiment. The x axis represents the position (in bp) on the phage genome.
- Extended Data Figure 8: Infection experiments with phi3T ΔaimP and derived conditioned media. (211 KB)
a–c, Growth curves of B. subtilis 168 infected with phi3T or phi3T ΔaimP in conditioned medium derived from phi3T (a); conditioned medium derived from phi3T ΔaimP (b); and control medium (c). Each medium was generated in biological replicate, and the two replicates are presented separately. Each curve represents the average of three technical replicates, and error bars represent s.e.
Extended Data Tables
- Supplementary Table 1 (187 KB)
This file shows the Homologs of AimR-AimP in sequenced genomes. Positions of AimR homologs are provided, as well as the corresponding full pro-pre-peptide sequence and the mature peptide sequences.
- Supplementary Table 2 (38 KB)
This file contains the annotation of the phi3T genome.