CI:Mor interactions in the lysogeny switches of Lactococcus lactis TP901-1 and Staphylococcus aureus φ13 bacteriophages

Abstract

Aim: To structurally characterize in detail the interactions between the phage repressor (CI) and the antirepressor (Mor) in the lysis-lysogeny switches of two Gram-positive bacteriophages, the lactococcal TP901-1 and staphylococcal φ13.

Methods: We use crystallographic structure determination, computational structural modeling, and analysis, as well as biochemical methods, to elucidate similarities and differences in the CI:Mor interactions for the two genetic switches.

Results: By comparing a newly determined and other available crystal structures for the N-terminal domain of CI (CI-NTD), we show that the CI interface involved in Mor binding undergoes structural changes upon binding in TP901-1. Most importantly, we show experimentally for the first time the direct interaction between CI and Mor for φ13, and model computationally the interaction interface. The computational modeling supports similar side chain rearrangements in TP901-1 and φ13.

Conclusion: This study ascertains experimentally that, like in the TP901-1 lysogeny switch, staphylococcal φ13 CI and Mor interact with each other. The structural basis of the interaction of φ13 CI and Mor was computationally modeled and is similar to the interaction demonstrated experimentally between TP901-1 CI-NTD and Mor, likely involving similar rearrangement of residue side chains during the formation of the complex. The study identifies one CI residue, Glu69, which unusually interacts primarily through its aliphatic chain with an aromatic residue on Mor after changing its conformation compared to the un-complexed structure. This and other residues at the interface are suggested for investigation in future studies.

Keywords

Lysogeny switch / temperate phage / repressor / antirepressor / corepressor / pathogen / human adaptation

References

[1] Turner NA,Maskarinec SA.Methicillin-resistant Staphylococcus aureus: an overview of basic and clinical research.Nat Rev Microbiol2019;17:203-18 PMCID:PMC6939889

[2] Lindsay JA.Understanding the rise of the superbug: investigation of the evolution and genomic variation of Staphylococcus aureus.Funct Integr Genomics2006;6:186-201

[3] McGuinness WA,DeLeo FR.Vancomycin resistance in Staphylococcus aureus.Yale J Biol Med2017;90:269-81 PMCID:PMC5482303

[4] Brady A,Gallego del Sol F,Quiles-Puchalt N.Molecular basis of lysis-lysogeny decisions in gram-positive phages.Annu Rev Microbiol2021;75:563-81

[5] Haaber J,Cohn MT.Bacterial viruses enable their host to acquire antibiotic resistance genes from neighbouring cells.Nat Commun2016;7:13333 PMCID:PMC5103068

[6] Ingmer H,Wolz C.Temperate phages of Staphylococcus aureus.Microbiol Spectr2019;7:10-1128

[7] Goerke C,Holtfreter S.Diversity of prophages in dominant Staphylococcus aureus clonal lineages.J Bacteriol2009;191:3462-8 PMCID:PMC2681900

[8] Rohmer C.The role of hlb-converting bacteriophages in Staphylococcus aureus host adaption.Microb Physiol2021;31:109-22

[9] de Jong NWM, van Kessel KPM, van Strijp JAG. Immune evasion by Staphylococcus aureus.Microbiol Spectr2019;7

[10] Sieber RN,Petersen A.Phage-mediated immune evasion and transmission of livestock-associated methicillin-resistant Staphylococcus aureus in humans.Emerg Infect Dis2020;26:2578-85 PMCID:PMC7588543

[11] Verkade E.Livestock-associated Staphylococcus aureus CC398: animal reservoirs and human infections.Infect Genet Evol2014;21:523-30

[12] Leinweber H,Larsen J,Ingmer H.Staphylococcal phages adapt to new hosts by extensive attachment site variability.mBio2021;12:e0225921 PMCID:PMC8649754

[13] Chevallereau A,van Houte S.Interactions between bacterial and phage communities in natural environments.Nat Rev Microbiol2022;20:49-62

[14] Gordillo Altamirano FL, Barr JJ. Phage therapy in the postantibiotic era.Clin Microbiol Rev2019;32:e00066-18 PMCID:PMC6431132

[15] Ptashne M.Principles of a switch.Nat Chem Biol2011;7:484-7

[16] Galkin VE,Bielnicki J,Bell CE.Cleavage of bacteriophage lambda cI repressor involves the RecA C-terminal domain.J Mol Biol2009;385:779-87 PMCID:PMC2648975

[17] Roberts JW.Proteolytic cleavage of bacteriophage lambda repressor in induction.Proc Natl Acad Sci U S A1975;72:147-51 PMCID:PMC432259

[18] Pedersen M,Grossmann JG,Hammer K.Identification of quaternary structure and functional domains of the CI repressor from bacteriophage TP901-1.J Mol Biol2008;376:983-96

[19] Frandsen KH,Jensen MR.Binding of the N-terminal domain of the lactococcal bacteriophage TP901-1 CI repressor to its target DNA: a crystallography, small angle scattering, and nuclear magnetic resonance study.Biochemistry2013;52:6892-904

[20] Rasmussen KK,Boeri Erba EB.Structural and dynamics studies of a truncated variant of CI repressor from bacteriophage TP901-1.Sci Rep2016;6:29574 PMCID:PMC4941734

[21] Pedersen M,Cassias J.Repression of the lysogenic P_R promoter in bacteriophage TP901-1 through binding of a CI-MOR complex to a composite O_M-O_R operator.Sci Rep2020;10:8659 PMCID:PMC7250872

[22] Varming AK,Zong Z,Kilstrup M.Flexible linker modulates the binding affinity of the TP901-1 CI phage repressor to DNA.FEBS J2022;289:1135-48

[23] Rasmussen KK,Schmidt SN.Structural basis of the bacteriophage TP901-1 CI repressor dimerization and interaction with DNA.FEBS Lett2018;592:1738-50

[24] Rasmussen KK,Varming AK.Revealing the mechanism of repressor inactivation during switching of a temperate bacteriophage.Proc Natl Acad Sci U S A2020;117:20576-85 PMCID:PMC7456139

[25] Madsen PL,Hammer K.The genetic switch regulating activity of early promoters of the temperate lactococcal bacteriophage TP901-1.J Bacteriol1999;181:7430-8 PMCID:PMC94198

[26] Das A,Hemmadi V,Biswas M.Studies on the gene regulation involved in the lytic-lysogenic switch in Staphylococcus aureus temperate bacteriophage Phi11.J Biochem2020;168:659-68

[27] Biswas A,Sau S.The N-terminal domain of the repressor of Staphylococcus aureus phage Φ11 possesses an unusual dimerization ability and DNA binding affinity.PLoS One2014;9:e95012 PMCID:PMC3991615

[28] Kristensen CS,Leinweber HAK.Characterization of the genetic switch from phage ɸ13 important for Staphylococcus aureus colonization in humans.Microbiologyopen2021;10:e1245 PMCID:PMC8516035

[29] Tang Y,Hvitved A.Commercial biocides induce transfer of prophage Φ13 from human strains of Staphylococcus aureus to livestock CC398.Front Microbiol2017;8:2418 PMCID:PMC5726172

[30] Winn MD,Cowtan KD.Overview of the CCP4 suite and current developments.Acta Crystallogr D Biol Crystallogr2011;67:235-42 PMCID:PMC3069738

[31] Vagin A.MOLREP : an automated program for molecular replacement.J Appl Crystallogr1997;30:1022-5

[32] Vagin AA,Lebedev AA.REFMAC5 dictionary: organization of prior chemical knowledge and guidelines for its use.Acta Crystallogr D Biol Crystallogr2004;60:2184-95

[33] Emsley P,Scott WG.Features and development of Coot.Acta Crystallogr D Biol Crystallogr2010;66:486-501 PMCID:PMC2852313

[34] Lovell SC,Arendall WB III.Structure validation by Cα geometry: ϕ, ψ and Cβ deviation.Proteins2003;50:437-50

[35] Varadi M,Deshpande M.AlphaFold Protein Structure Database: massively expanding the structural coverage of protein-sequence space with high-accuracy models.Nucleic Acids Res2022;50:D439-44 PMCID:PMC8728224

[36] Mirdita M,Moriwaki Y,Ovchinnikov S.ColabFold: making protein folding accessible to all.Nat Methods2022;19:679-82 PMCID:PMC9184281

[37] Rohmer C,Tuncbilek-Dere D.Influence of Staphylococcus aureus strain background on Sa3int phage life cycle switches.Viruses2022;14:2471 PMCID:PMC9694928

[38] Rezaei Javan R, Ramos-Sevillano E, Akter A, Brown J, Brueggemann AB. Prophages and satellite prophages are widespread in Streptococcus and may play a role in pneumococcal pathogenesis.Nat Commun2019;10:4852 PMCID:PMC6813308

[39] Matos RC,Rigottier-Gois L.Enterococcus faecalis prophage dynamics and contributions to pathogenic traits.PLoS Genet2013;9:e1003539 PMCID:PMC3675006