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Segrosome structure revealed by a complex of ParR with centromere DNA

Abstract

The stable inheritance of genetic material depends on accurate DNA partition. Plasmids serve as tractable model systems to study DNA segregation because they require only a DNA centromere, a centromere-binding protein and a force-generating ATPase. The centromeres of partition (par) systems typically consist of a tandem arrangement of direct repeats1,2,3,4,5,6,7. The best-characterized par system contains a centromere-binding protein called ParR and an ATPase called ParM. In the first step of segregation, multiple ParR proteins interact with the centromere repeats to form a large nucleoprotein complex of unknown structure called the segrosome, which binds ParM filaments4,8,9,10. pSK41 ParR binds a centromere consisting of multiple 20-base-pair (bp) tandem repeats to mediate both transcription autoregulation and segregation. Here we report the structure of the pSK41 segrosome revealed in the crystal structure of a ParR–DNA complex. In the crystals, the 20-mer tandem repeats stack pseudo-continuously to generate the full-length centromere with the ribbon–helix–helix (RHH) fold of ParR binding successive DNA repeats as dimer-of-dimers. Remarkably, the dimer-of-dimers assemble in a continuous protein super-helical array, wrapping the DNA about its positive convex surface to form a large segrosome with an open, solenoid-shaped structure, suggesting a mechanism for ParM capture and subsequent plasmid segregation.

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Figure 1: The pSK41 centromere-like site.
Figure 2: Structure of pSK41 ParR–DNA segrosome.
Figure 3: ParR–DNA and ParR–ParR interactions in the segrosome.
Figure 4: pSK41 segregation model.

References

  1. Møller-Jensen, J., Jensen, R. B. & Gerdes, K. Plasmid and chromosome segregation in prokaryotes. Trends Microbiol. 8, 313–320 (2000)

    Article  Google Scholar 

  2. Schumacher, M. A. Structural biology of plasmid segregation proteins. Curr. Opin. Struct. Biol. 17, 103–107 (2007)

    CAS  Article  Google Scholar 

  3. Gerdes, K., Møller-Jensen, J., Ebersbach, G., Kruse, T. & Nordström, K. Bacterial mitotic machineries. Cell 116, 359–366 (2004)

    CAS  Article  Google Scholar 

  4. Hayes, F. & Barillà, D. The bacterial segrosome: a dynamic nucleoprotein machine for DNA trafficking and segregation. Nat. Rev. Microbiol. 4, 133–143 (2006)

    CAS  Article  Google Scholar 

  5. Jensen, R. B. & Gerdes, K. Partitioning of plasmid R1. The ParM protein exhibits ATPase activity and interacts with the centromere-like ParR-parC complex. J. Mol. Biol. 269, 505–513 (1997)

    CAS  Article  Google Scholar 

  6. Jensen, R. B., Lurz, R. & Gerdes, K. Mechanism of DNA segregation in prokaryotes: replicon pairing by parC of plasmid R1. Proc. Natl Acad. Sci. USA 95, 8550–8555 (1998)

    ADS  CAS  Article  Google Scholar 

  7. Dam, M. & Gerdes, K. Partitioning of plasmid R1. Ten direct repeats flanking the parA promoter constitute a centromere-like partitioning site parC, that expresses incompatibility. J. Mol. Biol. 236, 1289–1298 (1994)

    CAS  Article  Google Scholar 

  8. Garner, E. C., Campbell, C. S., Weibel, D. B. & Mullins, R. D. Reconstitution of DNA segregation driven by assembly of a prokaryotic actin homolog. Science 315, 1270–1274 (2007)

    ADS  CAS  Article  Google Scholar 

  9. Møller-Jensen, J., Jensen, R. B., Löwe, J. & Gerdes, K. Prokaryotic DNA segregation by an actin-like filament. EMBO J. 21, 3119–3127 (2002)

    Article  Google Scholar 

  10. Møller-Jensen, J. et al. Bacterial mitosis: ParM of plasmid R1 moves plasmid DNA by an actin-like insertional polymerization mechanism. Mol. Cell 12, 1477–1487 (2003)

    Article  Google Scholar 

  11. Larsen, R. A. et al. Treadmilling of a prokaryotic tubulin-like protein, TubZ, required for plasmid stability. Genes Dev. 21, 1340–1352 (2007)

    CAS  Article  Google Scholar 

  12. Lundblad, J. R., Laurance, M. & Goodman, R. H. Fluorescence polarization of protein–DNA and protein–protein interactions. J. Mol. Endocrinol. 10, 607–612 (1996)

    CAS  Google Scholar 

  13. Terwilliger, T. C. & Berendzen, J. Automated MAD and MIR structure solution. Acta Crystallogr. D 55, 849–861 (1999)

    CAS  Article  Google Scholar 

  14. Brünger, A. T. et al. Crystallography and NMR System: a new software suite for macromolecular structure determination. Acta Crystallogr. D 54, 905–921 (1998)

    Article  Google Scholar 

  15. Raumann, B. E., Rould, M. A., Pabo, C. O. & Sauer, R. T. DNA recognition by β-sheets in the Arc repressor–operator crystal structure. Nature 367, 754–757 (1994a)

    ADS  CAS  Article  Google Scholar 

  16. Raumann, B. E., Brown, B. M. & Sauer, R. T. Major groove DNA recognition by β-sheets: the ribbon–helix–helix family of gene regulatory proteins. Curr. Opin. Struct. Biol. 4, 36–43 (1994b)

    CAS  Article  Google Scholar 

  17. Weihofen, W. A., Cicek, A., Pratto, F., Alonso, J. C. & Saenger, W. Structures of ω repressors bound to direct and inverted DNA repeats explain modulation of transcription. Nucleic Acids Res. 34, 1450–1458 (2006)

    CAS  Article  Google Scholar 

  18. Somers, W. S. & Phillips, S. E. Crystal structure of the met repressor–operator complex at 2.8 Å resolution reveals DNA recognition by β-strands. Nature 359, 387–393 (1992)

    ADS  CAS  Article  Google Scholar 

  19. He, Y. Y. et al. Probing the met repressor–operator recognition in solution. Nature 359, 431–433 (1992)

    ADS  CAS  Article  Google Scholar 

  20. Larson, J. D. et al. Crystal structures of the DNA-binding domain of Escherichia coli proline utilization A flavoprotein and analysis of the role of Lys9 in DNA recognition. Protein Sci. 15, 2630–2641 (2006)

    CAS  Article  Google Scholar 

  21. Golovanov, A. P., Barillà, D., Golovanova, M., Hayes, F. & Lian, L. Y. ParG, a protein required for active partition of bacterial plasmids, has a dimeric ribbon–helix–helix structure. Mol. Microbiol. 50, 1141–1153 (2003)

    CAS  Article  Google Scholar 

  22. Luger, K., Mäder, A., Richmond, R. K., Sargent, D. F. & Richmond, T. J. Crystal structure of the nucleosome core particle at 2.8 Å resolution. Nature 389, 251–260 (1997)

    ADS  CAS  Article  Google Scholar 

  23. Ravishanker, G., Swaminathan, S., Beveridge, D. L., Lavery, R. & Sklenar, H. Conformational and helicoidal analysis of 30 PS of molecular dynamics on the d(CGCGAATTCGCG) double helix: ‘curves’, dials and windows. J. Biomol. Struct. Dyn. 6, 669–699 (1998)

    Article  Google Scholar 

  24. Zhang, Y., Xi, Z., Hedge, R. S., Shakked, Z. & Crothers, D. M. Predicting indirect readout effects in protein–DNA interactions. Proc. Natl Acad. Sci. USA 101, 8337–8341 (2004)

    ADS  CAS  Article  Google Scholar 

  25. Rodionov, O., Lobocka, M. & Yarmolinsky, M. Silencing of genes flanking the P1 plasmid centromere. Science 283, 546–549 (1999)

    ADS  CAS  Article  Google Scholar 

  26. Schumacher, M. A. & Funnell, B. E. Structures of ParB bound to DNA reveal mechanism of partition complex formation. Nature 438, 516–519 (2005)

    ADS  CAS  Article  Google Scholar 

  27. Khare, D., Ziegelin, G., Lanka, E. & Heinemann, U. Sequence-specific DNA binding determined by contacts outside the helix–turn–helix motif of the ParB homolog KorB. Nat. Struct. Mol. Biol. 11, 656–663 (2004)

    CAS  Article  Google Scholar 

  28. Delano, W. L. The PyMOL Molecular Graphics System (DeLano Scientific, San Carlos, California, 2002)

    Google Scholar 

  29. Nicholls, A., Sharp, K. A. & Honig, B. Protein folding and association: insights from the interfacial and thermodynamic properties of hydrocarbons. Proteins 11, 281–296 (1991)

    CAS  Article  Google Scholar 

  30. van den Ent, F., Møller-Jensen, J., Amos, L. A., Gerdes, K. & Löwe, J. F-actin-like filaments formed by plasmid segregation protein ParM. EMBO J. 21, 6935–6943 (2002)

    CAS  Article  Google Scholar 

  31. Kabsch, W. & Holmes, K. C. The actin fold. FASEB J. 9, 167–174 (1995)

    CAS  Article  Google Scholar 

  32. Bork, P., Sander, C. & Valencia, A. An ATPase domain common to prokaryotic cell cycle proteins, sugar kinases, actin and hsp70 heat shock proteins. Proc. Natl Acad. Sci. USA 89, 7290–7294 (1992)

    ADS  CAS  Article  Google Scholar 

  33. Kwong, S. M., Skurray, R. A. & Firth, N. Staphylococcus aureus multiresistance plasmid pSK41: analysis of the replication region, initiator protein binding and antisense RNA regulation. Mol. Microbiol. 51, 497–509 (2004)

    CAS  Article  Google Scholar 

  34. Jones, T. A., Zou, J.-Y., Cowan, S. W. & Kjeldgaard, M. Improved methods for building protein models in electron density maps and the location of errors in these models. Acta Crystallogr. A 47, 110–119 (1991)

    Article  Google Scholar 

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Acknowledgements

We thank the Advanced Light Source (ALS) and their support staff. The ALS is supported by the Director, Office of Science, Office of Basic Energy Sciences, Materials Sciences Division of the US Department of Energy at the Lawrence Berkeley National Laboratory. We also thank Dr S. Kwong for supplying shuttle vectors and Professor K. Gerdes for pointing out the presence of the pSK41 par system. This work was supported by a Burroughs Wellcome Career Development Award, a U.T. M.D. Anderson Trust Fellowship and a National Institutes of Health grant (to M.A.S.), an Australian Research Council Grant (to N.F. and R.A.S.) and a National Health and Medical Research Council (Australia) Project Grant (to R.A.S. and N.F.).

Author Contributions M.A.S. performed the crystallographic studies, fluorescence polarization, oversaw cryo-electron microscopy studies and wrote the manuscript. T.C.G. generated ParRN and ParRC constructs. T.D.D. performed cryo-electron microscopy studies. A.J.B. generated plasmid constructs and undertook the regulatory and functional studies. S.O.J. performed the electrophoretic mobility shift assays (EMSA) and footprinting assays and contributed to the manuscript. R.A.S. and N.F. conceived and oversaw the functional studies and contributed to the manuscript.

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Correspondence to Maria A. Schumacher.

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Supplementary Information

The file contains Supplementary Discussion describing the clinical relevance of the pSK41 par system. This section also provides more in depth discussion regarding the CAT assays, DNase I protection and cryo-EM studies. Also included, there are Supplementary Figures 1-6 with Legends and Supplementary Tables 1-2. (PDF 1109 kb)

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Schumacher, M., Glover, T., Brzoska, A. et al. Segrosome structure revealed by a complex of ParR with centromere DNA. Nature 450, 1268–1271 (2007). https://doi.org/10.1038/nature06392

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