Organization of peptidoglycan synthesis in nodes and separate rings at different stages of cell division of Streptococcus pneumoniae

Amilcar J. Perez; Michael J. Boersma; Kevin E. Bruce; Melissa M. Lamanna; Sidney L. Shaw; Ho Ching T. Tsui; Atsushi Taguchi; Erin E. Carlson; Michael S. VanNieuwenhze; Malcolm E. Winkler

doi:10.1111/mmi.14659

Organization of peptidoglycan synthesis in nodes and separate rings at different stages of cell division of Streptococcus pneumoniae

Amilcar J. Perez, Michael J. Boersma, Kevin E. Bruce, Melissa M. Lamanna, Sidney L. Shaw, Ho Ching T. Tsui, Atsushi Taguchi, Erin E. Carlson, Michael S. VanNieuwenhze, Malcolm E. Winkler

Chemistry (Twin Cities)

Research output: Contribution to journal › Article › peer-review

22 Scopus citations

Abstract

Bacterial peptidoglycan (PG) synthesis requires strict spatiotemporal organization to reproduce specific cell shapes. In ovoid-shaped Streptococcus pneumoniae (Spn), septal and peripheral (elongation) PG synthesis occur simultaneously at midcell. To uncover the organization of proteins and activities that carry out these two modes of PG synthesis, we examined Spn cells vertically oriented onto their poles to image the division plane at the high lateral resolution of 3D-SIM (structured-illumination microscopy). Labeling with fluorescent D-amino acids (FDAA) showed that areas of new transpeptidase (TP) activity catalyzed by penicillin-binding proteins (PBPs) separate into a pair of concentric rings early in division, representing peripheral PG (pPG) synthesis (outer ring) and the leading-edge (inner ring) of septal PG (sPG) synthesis. Fluorescently tagged PBP2x or FtsZ locate primarily to the inner FDAA-marked ring, whereas PBP2b and FtsX remain in the outer ring, suggesting roles in sPG or pPG synthesis, respectively. Pulses of FDAA labeling revealed an arrangement of separate regularly spaced “nodes” of TP activity around the division site of predivisional cells. Tagged PBP2x, PBP2b, and FtsX proteins also exhibited nodal patterns with spacing comparable to that of FDAA labeling. Together, these results reveal new aspects of spatially ordered PG synthesis in ovococcal bacteria during cell division.

Original language	English (US)
Pages (from-to)	1152-1169
Number of pages	18
Journal	Molecular Microbiology
Volume	115
Issue number	6
Early online date	Dec 2 2020
DOIs	https://doi.org/10.1111/mmi.14659
State	Published - Jun 2021

Bibliographical note

Funding Information:
We thank laboratory members and Erkin Kuru (Harvard Med Sch), Yves Brun (Universit? de Montr?al), Seamus Holden (Newcastle Univ), and Jie Xiao and Josh McCausland (Johns Hopkins) for discussions and suggestions; Jim Powers (IUB) for help with microscopy, Luke Lavis (Janelia Lab) for Fluor JF549; Dalia Denapaite (Trento Univ), Reinhold Br?ckner, and Regine Hakenbeck (Kaiserlautern Univ) for anti-bPBP2x antibody; and Suzanne Walker and David Z. Rudner (Harvard Med Sch) for supporting preparation of anti-bPBP2x and anti-bPBP2b antibodies. This work was supported by NIH Grants RO1GM113172 (to MSV and MEW); RO1GM128439 (to EEC and MEW); R35GM131767 (to MEW); R35GM136365 (to MSV), RO1AI148752 (to S. Walker), RO1AI139083 (To D. Z. Rudner), National Science Foundation Grant MCB1615907 (to S.L.S.); NIH Predoctoral Quantitative and Chemical Biology Training Grant T32 GM109825 (to AJP); NIH Predoctoral Grant F31AI138430 (to MML); and NIH Equipment Grant S10OD024988 to the Indiana University Bloomington (IUB) Light Microscopy Imaging Center.

Funding Information:
We thank laboratory members and Erkin Kuru (Harvard Med Sch), Yves Brun (Université de Montréal), Seamus Holden (Newcastle Univ), and Jie Xiao and Josh McCausland (Johns Hopkins) for discussions and suggestions; Jim Powers (IUB) for help with microscopy, Luke Lavis (Janelia Lab) for Fluor JF549; Dalia Denapaite (Trento Univ), Reinhold Brückner, and Regine Hakenbeck (Kaiserlautern Univ) for anti‐bPBP2x antibody; and Suzanne Walker and David Z. Rudner (Harvard Med Sch) for supporting preparation of anti‐bPBP2x and anti‐bPBP2b antibodies. This work was supported by NIH Grants RO1GM113172 (to MSV and MEW); RO1GM128439 (to EEC and MEW); R35GM131767 (to MEW); R35GM136365 (to MSV), RO1AI148752 (to S. Walker), RO1AI139083 (To D. Z. Rudner), National Science Foundation Grant MCB1615907 (to S.L.S.); NIH Predoctoral Quantitative and Chemical Biology Training Grant T32 GM109825 (to AJP); NIH Predoctoral Grant F31AI138430 (to MML); and NIH Equipment Grant S10OD024988 to the Indiana University Bloomington (IUB) Light Microscopy Imaging Center.

Publisher Copyright:
© 2020 John Wiley & Sons Ltd

Keywords

FtsX localization
FtsZ localization
fluorescent D-amino acids (FDAAs)
penicillin-binding proteins (PBPs)
peripheral (elongation) peptidoglycan synthesis
septal peptidoglycan synthesis

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1111/mmi.14659

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Perez, A. J., Boersma, M. J., Bruce, K. E., Lamanna, M. M., Shaw, S. L., Tsui, H. C. T., Taguchi, A., Carlson, E. E., VanNieuwenhze, M. S., & Winkler, M. E. (2021). Organization of peptidoglycan synthesis in nodes and separate rings at different stages of cell division of Streptococcus pneumoniae. Molecular Microbiology, 115(6), 1152-1169. https://doi.org/10.1111/mmi.14659

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title = "Organization of peptidoglycan synthesis in nodes and separate rings at different stages of cell division of Streptococcus pneumoniae",

abstract = "Bacterial peptidoglycan (PG) synthesis requires strict spatiotemporal organization to reproduce specific cell shapes. In ovoid-shaped Streptococcus pneumoniae (Spn), septal and peripheral (elongation) PG synthesis occur simultaneously at midcell. To uncover the organization of proteins and activities that carry out these two modes of PG synthesis, we examined Spn cells vertically oriented onto their poles to image the division plane at the high lateral resolution of 3D-SIM (structured-illumination microscopy). Labeling with fluorescent D-amino acids (FDAA) showed that areas of new transpeptidase (TP) activity catalyzed by penicillin-binding proteins (PBPs) separate into a pair of concentric rings early in division, representing peripheral PG (pPG) synthesis (outer ring) and the leading-edge (inner ring) of septal PG (sPG) synthesis. Fluorescently tagged PBP2x or FtsZ locate primarily to the inner FDAA-marked ring, whereas PBP2b and FtsX remain in the outer ring, suggesting roles in sPG or pPG synthesis, respectively. Pulses of FDAA labeling revealed an arrangement of separate regularly spaced “nodes” of TP activity around the division site of predivisional cells. Tagged PBP2x, PBP2b, and FtsX proteins also exhibited nodal patterns with spacing comparable to that of FDAA labeling. Together, these results reveal new aspects of spatially ordered PG synthesis in ovococcal bacteria during cell division.",

keywords = "FtsX localization, FtsZ localization, fluorescent D-amino acids (FDAAs), penicillin-binding proteins (PBPs), peripheral (elongation) peptidoglycan synthesis, septal peptidoglycan synthesis",

author = "Perez, {Amilcar J.} and Boersma, {Michael J.} and Bruce, {Kevin E.} and Lamanna, {Melissa M.} and Shaw, {Sidney L.} and Tsui, {Ho Ching T.} and Atsushi Taguchi and Carlson, {Erin E.} and VanNieuwenhze, {Michael S.} and Winkler, {Malcolm E.}",

note = "Funding Information: We thank laboratory members and Erkin Kuru (Harvard Med Sch), Yves Brun (Universit? de Montr?al), Seamus Holden (Newcastle Univ), and Jie Xiao and Josh McCausland (Johns Hopkins) for discussions and suggestions; Jim Powers (IUB) for help with microscopy, Luke Lavis (Janelia Lab) for Fluor JF549; Dalia Denapaite (Trento Univ), Reinhold Br?ckner, and Regine Hakenbeck (Kaiserlautern Univ) for anti-bPBP2x antibody; and Suzanne Walker and David Z. Rudner (Harvard Med Sch) for supporting preparation of anti-bPBP2x and anti-bPBP2b antibodies. This work was supported by NIH Grants RO1GM113172 (to MSV and MEW); RO1GM128439 (to EEC and MEW); R35GM131767 (to MEW); R35GM136365 (to MSV), RO1AI148752 (to S. Walker), RO1AI139083 (To D. Z. Rudner), National Science Foundation Grant MCB1615907 (to S.L.S.); NIH Predoctoral Quantitative and Chemical Biology Training Grant T32 GM109825 (to AJP); NIH Predoctoral Grant F31AI138430 (to MML); and NIH Equipment Grant S10OD024988 to the Indiana University Bloomington (IUB) Light Microscopy Imaging Center. Funding Information: We thank laboratory members and Erkin Kuru (Harvard Med Sch), Yves Brun (Universit{\'e} de Montr{\'e}al), Seamus Holden (Newcastle Univ), and Jie Xiao and Josh McCausland (Johns Hopkins) for discussions and suggestions; Jim Powers (IUB) for help with microscopy, Luke Lavis (Janelia Lab) for Fluor JF549; Dalia Denapaite (Trento Univ), Reinhold Br{\"u}ckner, and Regine Hakenbeck (Kaiserlautern Univ) for anti‐bPBP2x antibody; and Suzanne Walker and David Z. Rudner (Harvard Med Sch) for supporting preparation of anti‐bPBP2x and anti‐bPBP2b antibodies. This work was supported by NIH Grants RO1GM113172 (to MSV and MEW); RO1GM128439 (to EEC and MEW); R35GM131767 (to MEW); R35GM136365 (to MSV), RO1AI148752 (to S. Walker), RO1AI139083 (To D. Z. Rudner), National Science Foundation Grant MCB1615907 (to S.L.S.); NIH Predoctoral Quantitative and Chemical Biology Training Grant T32 GM109825 (to AJP); NIH Predoctoral Grant F31AI138430 (to MML); and NIH Equipment Grant S10OD024988 to the Indiana University Bloomington (IUB) Light Microscopy Imaging Center. Publisher Copyright: {\textcopyright} 2020 John Wiley & Sons Ltd",

year = "2021",

month = jun,

doi = "10.1111/mmi.14659",

language = "English (US)",

volume = "115",

pages = "1152--1169",

journal = "Molecular Microbiology",

issn = "0950-382X",

publisher = "John Wiley and Sons Inc.",

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T1 - Organization of peptidoglycan synthesis in nodes and separate rings at different stages of cell division of Streptococcus pneumoniae

AU - Perez, Amilcar J.

AU - Boersma, Michael J.

AU - Bruce, Kevin E.

AU - Lamanna, Melissa M.

AU - Shaw, Sidney L.

AU - Tsui, Ho Ching T.

AU - Taguchi, Atsushi

AU - Carlson, Erin E.

AU - VanNieuwenhze, Michael S.

AU - Winkler, Malcolm E.

N1 - Funding Information: We thank laboratory members and Erkin Kuru (Harvard Med Sch), Yves Brun (Universit? de Montr?al), Seamus Holden (Newcastle Univ), and Jie Xiao and Josh McCausland (Johns Hopkins) for discussions and suggestions; Jim Powers (IUB) for help with microscopy, Luke Lavis (Janelia Lab) for Fluor JF549; Dalia Denapaite (Trento Univ), Reinhold Br?ckner, and Regine Hakenbeck (Kaiserlautern Univ) for anti-bPBP2x antibody; and Suzanne Walker and David Z. Rudner (Harvard Med Sch) for supporting preparation of anti-bPBP2x and anti-bPBP2b antibodies. This work was supported by NIH Grants RO1GM113172 (to MSV and MEW); RO1GM128439 (to EEC and MEW); R35GM131767 (to MEW); R35GM136365 (to MSV), RO1AI148752 (to S. Walker), RO1AI139083 (To D. Z. Rudner), National Science Foundation Grant MCB1615907 (to S.L.S.); NIH Predoctoral Quantitative and Chemical Biology Training Grant T32 GM109825 (to AJP); NIH Predoctoral Grant F31AI138430 (to MML); and NIH Equipment Grant S10OD024988 to the Indiana University Bloomington (IUB) Light Microscopy Imaging Center. Funding Information: We thank laboratory members and Erkin Kuru (Harvard Med Sch), Yves Brun (Université de Montréal), Seamus Holden (Newcastle Univ), and Jie Xiao and Josh McCausland (Johns Hopkins) for discussions and suggestions; Jim Powers (IUB) for help with microscopy, Luke Lavis (Janelia Lab) for Fluor JF549; Dalia Denapaite (Trento Univ), Reinhold Brückner, and Regine Hakenbeck (Kaiserlautern Univ) for anti‐bPBP2x antibody; and Suzanne Walker and David Z. Rudner (Harvard Med Sch) for supporting preparation of anti‐bPBP2x and anti‐bPBP2b antibodies. This work was supported by NIH Grants RO1GM113172 (to MSV and MEW); RO1GM128439 (to EEC and MEW); R35GM131767 (to MEW); R35GM136365 (to MSV), RO1AI148752 (to S. Walker), RO1AI139083 (To D. Z. Rudner), National Science Foundation Grant MCB1615907 (to S.L.S.); NIH Predoctoral Quantitative and Chemical Biology Training Grant T32 GM109825 (to AJP); NIH Predoctoral Grant F31AI138430 (to MML); and NIH Equipment Grant S10OD024988 to the Indiana University Bloomington (IUB) Light Microscopy Imaging Center. Publisher Copyright: © 2020 John Wiley & Sons Ltd

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N2 - Bacterial peptidoglycan (PG) synthesis requires strict spatiotemporal organization to reproduce specific cell shapes. In ovoid-shaped Streptococcus pneumoniae (Spn), septal and peripheral (elongation) PG synthesis occur simultaneously at midcell. To uncover the organization of proteins and activities that carry out these two modes of PG synthesis, we examined Spn cells vertically oriented onto their poles to image the division plane at the high lateral resolution of 3D-SIM (structured-illumination microscopy). Labeling with fluorescent D-amino acids (FDAA) showed that areas of new transpeptidase (TP) activity catalyzed by penicillin-binding proteins (PBPs) separate into a pair of concentric rings early in division, representing peripheral PG (pPG) synthesis (outer ring) and the leading-edge (inner ring) of septal PG (sPG) synthesis. Fluorescently tagged PBP2x or FtsZ locate primarily to the inner FDAA-marked ring, whereas PBP2b and FtsX remain in the outer ring, suggesting roles in sPG or pPG synthesis, respectively. Pulses of FDAA labeling revealed an arrangement of separate regularly spaced “nodes” of TP activity around the division site of predivisional cells. Tagged PBP2x, PBP2b, and FtsX proteins also exhibited nodal patterns with spacing comparable to that of FDAA labeling. Together, these results reveal new aspects of spatially ordered PG synthesis in ovococcal bacteria during cell division.

AB - Bacterial peptidoglycan (PG) synthesis requires strict spatiotemporal organization to reproduce specific cell shapes. In ovoid-shaped Streptococcus pneumoniae (Spn), septal and peripheral (elongation) PG synthesis occur simultaneously at midcell. To uncover the organization of proteins and activities that carry out these two modes of PG synthesis, we examined Spn cells vertically oriented onto their poles to image the division plane at the high lateral resolution of 3D-SIM (structured-illumination microscopy). Labeling with fluorescent D-amino acids (FDAA) showed that areas of new transpeptidase (TP) activity catalyzed by penicillin-binding proteins (PBPs) separate into a pair of concentric rings early in division, representing peripheral PG (pPG) synthesis (outer ring) and the leading-edge (inner ring) of septal PG (sPG) synthesis. Fluorescently tagged PBP2x or FtsZ locate primarily to the inner FDAA-marked ring, whereas PBP2b and FtsX remain in the outer ring, suggesting roles in sPG or pPG synthesis, respectively. Pulses of FDAA labeling revealed an arrangement of separate regularly spaced “nodes” of TP activity around the division site of predivisional cells. Tagged PBP2x, PBP2b, and FtsX proteins also exhibited nodal patterns with spacing comparable to that of FDAA labeling. Together, these results reveal new aspects of spatially ordered PG synthesis in ovococcal bacteria during cell division.

KW - FtsX localization

KW - FtsZ localization

KW - fluorescent D-amino acids (FDAAs)

KW - penicillin-binding proteins (PBPs)

KW - peripheral (elongation) peptidoglycan synthesis

KW - septal peptidoglycan synthesis

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JO - Molecular Microbiology

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Organization of peptidoglycan synthesis in nodes and separate rings at different stages of cell division of Streptococcus pneumoniae

Abstract

Bibliographical note

Keywords

UN SDGs

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