A Vibriophage Takes Antirepression to the Next Level
Authors: Leonardo F. Lemos Rocha, Melanie Blokesch
Journal: Cell Host & Microbe (2020)

Since its isolation by Esther Lederberg, phage lambda and its repressor protein CI have contributed substantially to the advancement of molecular biology. In this issue of Cell Host & Microbe, Silpe et al., 2020Silpe J.E. Bridges A.A. Huang X. Coronado D.R. Duddy O.P. Bassler B.L. Separating Functions of the Phage-Encoded Quorum-Sensing-Activated Antirepressor Qtip.Cell Host Microbe. 2020; 27 (this issue): 629-641Abstract Full Text Full Text PDF Scopus (21) Google Scholar characterize the antirepressor Qtip of Vibrio phage VP882, which through CI sequestration triggers a lytic switch. Since its isolation by Esther Lederberg, phage lambda and its repressor protein CI have contributed substantially to the advancement of molecular biology. In this issue of Cell Host & Microbe, Silpe et al., 2020Silpe J.E. Bridges A.A. Huang X. Coronado D.R. Duddy O.P. Bassler B.L. Separating Functions of the Phage-Encoded Quorum-Sensing-Activated Antirepressor Qtip.Cell Host Microbe. 2020; 27 (this issue): 629-641Abstract Full Text Full Text PDF Scopus (21) Google Scholar characterize the antirepressor Qtip of Vibrio phage VP882, which through CI sequestration triggers a lytic switch. Bacteria-infecting viruses, known as bacteriophages or phages, are genetic parasites that have developed numerous strategies to ensure their replication within a host cell. Not all phages, however, are instant killers of their host; instead, temperate phages have acquired the ability to undergo lysogeny, thereby ensuring vertical transmission from the mother cell to its offspring. The lysogenic lifestyle relies on the silencing of the majority of the phage-carried genes so that capsid proteins or lytic enzymes are not synthesized. Here, Silpe, Bridges, and colleagues investigate how lytic genes are derepressed in the Vibrio phage VP882 to trigger cell lysis (Silpe et al., 2020Silpe J.E. Bridges A.A. Huang X. Coronado D.R. Duddy O.P. Bassler B.L. Separating Functions of the Phage-Encoded Quorum-Sensing-Activated Antirepressor Qtip.Cell Host Microbe. 2020; 27 (this issue): 629-641Abstract Full Text Full Text PDF Scopus (21) Google Scholar). The best-studied temperate phage is without doubt phage λ. The isolation of λ by Esther Lederberg occurred 70 years ago and was based on a serendipitous observation (Lederberg, 1950Lederberg E.M. Lysogenicity in Escherichia coli strain K-12.Microbial Genetics Bulletin. 1950; 1: 5-8Google Scholar): she witnessed that Escherichia coli K-12 cells caused "nibbled and plaqued" growth of a sensitive E. coli strain W-518 when co-cultured. While many questions remained unanswered after this initial discovery, lambda biology is nowadays well understood (Casjens and Hendrix, 2015Casjens S.R. Hendrix R.W. Bacteriophage lambda: Early pioneer and still relevant.Virology. 2015; 479-480: 310-330Crossref PubMed Scopus (151) Google Scholar). Indeed, it is now appreciated that E. coli strain W-518 was a λ prophage-cured version of its parental strain (K-12) and that this curing resulted from earlier UV irradiation. The lysogenized parental strain itself remained immune to superinfection, and plaques were therefore absent from its monoculture. The molecular mechanisms that determine the lysogenic to lytic switch and the subsequent release of phage particles have been extensively studied. While the prophage is present in the host's genome, the cells keep producing the λ repressor protein CI, which silences other prophage genes, including those that drive cellular lysis. The lysogeny-lytic transition occurs stochastically at low frequencies within a bacterial population, with a more purposeful trigger occurring only when the host cell experiences stressful conditions that induce the bacterium's SOS response (Howard-Varona et al., 2017Howard-Varona C. Hargreaves K.R. Abedon S.T. Sullivan M.B. Lysogeny in nature: mechanisms, impact and ecology of temperate phages.ISME J. 2017; 11: 1511-1520Crossref PubMed Scopus (299) Google Scholar). One such stressor is UV irradiation, which, due to the DNA damage it causes, is frequently used to study bacterial SOS responses under laboratory settings. DNA damage leads to the appearance of single-stranded DNA (ssDNA), a substrate of the RecA protein (Baharoglu and Mazel, 2014Baharoglu Z. Mazel D. SOS, the formidable strategy of bacteria against aggressions.FEMS Microbiol. Rev. 2014; 38: 1126-1145Crossref PubMed Scopus (237) Google Scholar). In the presence of Mg2+ and ATP, RecA forms nucleofilaments on ssDNA; this activated form of RecA (RecA∗) triggers the auto-cleavage of the SOS repressor protein LexA. With repression released, the LexA regulon, which consists of about 40 genes in E. coli, is induced, with the ultimate aims being to alleviate the DNA damage and to overcome the stressor. As a structural homolog of LexA, the λ repressor CI likewise undergoes RecA∗-triggered self-cleavage and therefore loses its DNA binding ability, which ultimately abrogates phage gene repression and therefore induces the lytic cycle (Casjens and Hendrix, 2015Casjens S.R. Hendrix R.W. Bacteriophage lambda: Early pioneer and still relevant.Virology. 2015; 479-480: 310-330Crossref PubMed Scopus (151) Google Scholar). It is therefore obvious that the CI repressor is a master regulator for the lysogeny-lytic switch. Unsurprisingly, additional regulatory inputs into both the production and activity of CI repressor homologs have also evolved. For instance, numerous phages have adapted so called Arbitrium-like systems in which peptide-responsive non-coding RNAs control the expression of cI and therefore determine whether lysogeny is maintained or repealed (Erez et al., 2017Erez Z. Steinberger-Levy I. Shamir M. Doron S. Stokar-Avihail A. Peleg Y. Melamed S. Leavitt A. Savidor A. Albeck S. et al.Communication between viruses guides lysis-lysogeny decisions.Nature. 2017; 541: 488-493Crossref PubMed Scopus (317) Google Scholar, Stokar-Avihail et al., 2019Stokar-Avihail A. Tal N. Erez Z. Lopatina A. Sorek R. Widespread utilization of peptide communication in phages infecting soil and pathogenic bacteria.Cell Host Microbe. 2019; 25: 746-755.e745Abstract Full Text Full Text PDF PubMed Scopus (49) Google Scholar). Alternatively, CI repressor proteins can also be manipulated through the production of antirepressors. For example, the Salmonella phage P22 produces the antirepressor Ant, which inactivates CI proteins (named C2 in the case of P22) by sequestering them in a non-covalent manner, thereby preventing their binding to DNA (Susskind and Botstein, 1978Susskind M.M. Botstein D. Molecular genetics of bacteriophage P22.Microbiol. Rev. 1978; 42: 385-413Crossref PubMed Google Scholar). Interestingly, a further evolved variant of the antirepressor strategy was recently identified in the Vibrio parahaemolyticus phage VP882 (Silpe and Bassler, 2019Silpe J.E. Bassler B.L. A Host-Produced Quorum-Sensing Autoinducer Controls a Phage Lysis-Lysogeny Decision.Cell. 2019; 176: 268-280.e213Abstract Full Text Full Text PDF PubMed Scopus (151) Google Scholar). This "quorum-triggered inactivator of cI protein" (Qtip) antirepressor is only produced when a phage-encoded quorum sensing (QS) receptor senses the host-produced autoinducer DPO. Linking host lysis to a high-cell-density state has obvious advantages for phage propagation, as it implies an abundance of new host cells in the immediate surroundings. In this issue of Cell Host & Microbe, Silpe, Bridges, and colleagues further characterize this repressor-antirepressor interaction and identify regions and amino acids that distinguish them from the canonical CI and C2/Ant systems of λ and P22, respectively (Figure 1; Silpe et al., 2020Silpe J.E. Bridges A.A. Huang X. Coronado D.R. Duddy O.P. Bassler B.L. Separating Functions of the Phage-Encoded Quorum-Sensing-Activated Antirepressor Qtip.Cell Host Microbe. 2020; 27 (this issue): 629-641Abstract Full Text Full Text PDF Scopus (21) Google Scholar). The antirepressor Qtip is a small protein of 79 amino acids that recognizes the N-terminal domain of its cognate CI repressor (both from phage VP882) and thereby inhibits SOS-mediated autocleavage. Moreover, by reconstituting the system in vitro, the authors show that Qtip also prevents CI from binding to DNA. By combining in silico prediction with random, alanine-scanning, and site-directed mutagenesis of the CI and Qtip encoding genes, the authors identified key residues that are required for their interaction (Silpe et al., 2020Silpe J.E. Bridges A.A. Huang X. Coronado D.R. Duddy O.P. Bassler B.L. Separating Functions of the Phage-Encoded Quorum-Sensing-Activated Antirepressor Qtip.Cell Host Microbe. 2020; 27 (this issue): 629-641Abstract Full Text Full Text PDF Scopus (21) Google Scholar). Interestingly, the authors confirmed their previous observation that, in addition to CI binding, Qtip also sequesters its target protein and delocalizes it to the cell pole (Silpe and Bassler, 2019Silpe J.E. Bassler B.L. A Host-Produced Quorum-Sensing Autoinducer Controls a Phage Lysis-Lysogeny Decision.Cell. 2019; 176: 268-280.e213Abstract Full Text Full Text PDF PubMed Scopus (151) Google Scholar). Compared to the P22 antirepressor Ant, this feature appears unique to Qtip. Furthermore, while Qtip does not normally sequester unrelated CI proteins such as λ CI at the cell pole, polar sequestration could be restored in a chimera composed of the N-terminal domain of CIVP882 and the C-terminal domain of CIλ, although not vice versa. While the biological meaning of this polar localization was not investigated further in this study, this striking phenotype leaves room for speculation. For instance, one wonders whether the sequestration of CI compared to its SOS-induced autoproteolysis allows the bacterium to fine-tune and potentially invert the process under certain changing environmental conditions before the full lysis program is initiated. To test this hypothesis, one would need to look at the native bacterial host and its phage without artificial protein production. Indeed, as the authors acknowledged, natural Qtip levels are currently unknown, making a comparison between the induced protein level and the cell's native state impossible. Alternatively, could Qtip-induced polar aggregation interfere with the crucial processes that are controlled by the polar landmark protein HubP (Yamaichi et al., 2012Yamaichi Y. Bruckner R. Ringgaard S. Möll A. Cameron D.E. Briegel A. Jensen G.J. Davis B.M. Waldor M.K. A multidomain hub anchors the chromosome segregation and chemotactic machinery to the bacterial pole.Genes Dev. 2012; 26: 2348-2360Crossref PubMed Scopus (113) Google Scholar)? This protein plays an important role in chromosome segregation as well as in the assembly of the polar flagellum apparatus and the chemotaxis clusters. Indeed, the deletion of hubP results in a significant decrease in the bacterium's swarming activity and impairs chemotaxis (Yamaichi et al., 2012Yamaichi Y. Bruckner R. Ringgaard S. Möll A. Cameron D.E. Briegel A. Jensen G.J. Davis B.M. Waldor M.K. A multidomain hub anchors the chromosome segregation and chemotactic machinery to the bacterial pole.Genes Dev. 2012; 26: 2348-2360Crossref PubMed Scopus (113) Google Scholar). While not supported by any data at this point, it would be fascinating if the VP882 phage had not only evolved the ability to "listen in" on its host's QS system (Silpe and Bassler, 2019Silpe J.E. Bassler B.L. A Host-Produced Quorum-Sensing Autoinducer Controls a Phage Lysis-Lysogeny Decision.Cell. 2019; 176: 268-280.e213Abstract Full Text Full Text PDF PubMed Scopus (151) Google Scholar) but also to "paralyze" the host cell by interfering with flagellar assembly and chemotaxis. Such a state of paralysis could reduce the probability that the lysis-induced host could still go on to escape from the high-density environment before the released phages have had a chance to "hitch-hike" onto the neighboring bacteria. Undoubtedly, further exciting studies await that will expand our understanding of the fascinating mechanism of polar CI sequestration by the Qtip antirepressor protein. We acknowledge support by the European Research Council (consolidator grant 724630) and the Swiss National Science Foundation (grant 310030_185022). MB is a Howard Hughes Medical Institute (HHMI) International Research Scholar (grant 55008726). Separating Functions of the Phage-Encoded Quorum-Sensing-Activated Antirepressor QtipSilpe et al.Cell Host & MicrobeFebruary 25, 2020In BriefQtip is a 79 amino acid antirepressor that connects phage quorum sensing to host-cell lysis. Silpe and Bridges et al. demonstrate that Qtip binds, inactivates, and sequesters the phage cI repressor at the cell pole. Qtip mutagenesis reveals discrete sites required for repressor recognition and polar localization, showing that the two functions are separable. Full-Text PDF Open Archive