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PASTA repeats of the protein kinase StkP interconnect cell constriction and separation of Streptococcus pneumoniae

Nature Microbiology volume 3, pages 197–209 (2018)Cite this article

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Abstract

Eukaryotic-like serine/threonine kinases (eSTKs) with extracellular PASTA repeats are key membrane regulators of bacterial cell division. How PASTA repeats govern eSTK activation and function remains elusive. Using evolution- and structural-guided approaches combined with cell imaging, we disentangle the role of each PASTA repeat of the eSTK StkP from Streptococcus pneumoniae. While the three membrane-proximal PASTA repeats behave as interchangeable modules required for the activation of StkP independently of cell wall binding, they also control the septal cell wall thickness. In contrast, the fourth and membrane-distal PASTA repeat directs StkP localization at the division septum and encompasses a specific motif that is critical for final cell separation through interaction with the cell wall hydrolase LytB. We propose a model in which the extracellular four-PASTA domain of StkP plays a dual function in interconnecting the phosphorylation of StkP endogenous targets along with septal cell wall remodelling to allow cell division of the pneumococcus.

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Fig. 1: The PASTA4 repeat is critical for pneumococcal cell division.
Fig. 2: Deletion of PASTA1, 2 and 3 repeats inactivates StkP and renders pneumococcal cell division independent of StkP kinase activity.
Fig. 3: PASTA1, 2 and 3 are equivalent and required for StkP activation independently from their cell-wall binding specificity.
Fig. 4: PASTA4 localizes StkP at the division septum.
Fig. 5: Taxonomic distribution and organization of PASTA repeats in Streptococcaceae and structural organization of PASTA4 and its properties for cell division.
Fig. 6: PASTA4 positions LytB to the division septum while PASTA123 repeats rule the thickness of the septal cell wall.

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Acknowledgements

This work was supported by grants from the Centre National de la Recherche Scientifique, Université Claude Bernard Lyon 1, Agence Nationale de la Recherche (ANR-12-BSV3-0008-01 and ANR-15-CE32-0001-01), Region Auvergne-Rhône-Alpes (financial support for L.Z., C.M. and P.S.G.) and Bettencourt Schueller Foundation. We thank A. Ducret for implementing the heat maps and violin plots in MicrobeJ and carefully reading the paper. We thank P. Garcia at the Centro de Investigaciones Biológicas for providing the plasmid encoding GFP–LytB and the anti-LytB antibody, and L. S. Havarstein at the Norwegian University of Life Sciences for providing S. pneumoniae H130 genomic DNA. We acknowledge the contribution of the ‘Protein Science’ and ‘Plateau Technique Imagerie/Microscopie’ microscopy facilities of the Structure Fédérative de Recherche Biosciences Gerland-Lyon Sud (UMS344/US8), the ‘Centre Technologique des Microstructures’ of the Université Claude Bernard Lyon I, and Swiss Lightsource and European Synchrotron Radiation Facility synchrotron beamlines.

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Authors and Affiliations

  1. Molecular Microbiology and Structural Biochemistry, Unité Mixte de Recherche, Université Claude Bernard Lyon 1, Centre National de la Recherche Scientifique, Lyon, France

    Laure Zucchini, Chryslène Mercy, Pierre Simon Garcia, Frédéric Galisson, Céline Freton, Sébastien Guiral, Patrice Gouet & Christophe Grangeasse

  2. Laboratoire de Biométrie et Biologie Evolutive, Unité Mixte de Recherche, Université Claude Bernard Lyon 1, Centre National de la Recherche Scientifique, Villeurbanne, France

    Pierre Simon Garcia & Céline Brochier-Armanet

  3. Laboratoire de Biologie Tissulaire et d’Ingénierie Thérapeutique, Unité Mixte de Recherche, Université Claude Bernard Lyon 1, Centre National de la Recherche Scientifique, Lyon, France

    Caroline Cluzel

  4. Protein Science Facility, Structure Fédérative de Recherche Biosciences/UMS3444/US8, Université Claude Bernard Lyon 1, Ecole Normale Supérieur de Lyon, INSERM, Centre National de la Recherche Scientifique, Lyon, France

    Virginie Gueguen-Chaignon

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Contributions

L.Z., C.M. and C.F. conducted the cell biology and genetics experiments, purified proteins for the surface plasmon resonance experiments and performed the western blot analysis. F.G., P.G. and V.G.-C. conducted the crystallogenesis experiments and solved the structure of PASTA4. C.C. performed the transmission electron microscopy. P.S.G. and C.B-A. performed the phylogeny analyses. L.Z., C.M., P.S.G., C.C., V.G.-C., F.G., C.F. and S.G. designed and analysed the data with assistance from C.B.-A., P.G. and C.G. C.G. wrote the paper. All authors edited the paper.

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Correspondence to Christophe Grangeasse.

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Zucchini, L., Mercy, C., Garcia, P.S. et al. PASTA repeats of the protein kinase StkP interconnect cell constriction and separation of Streptococcus pneumoniae . Nat Microbiol 3, 197–209 (2018). https://doi.org/10.1038/s41564-017-0069-3

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