--- _id: '13116' abstract: - lang: eng text: 'The emergence of large-scale order in self-organized systems relies on local interactions between individual components. During bacterial cell division, FtsZ -- a prokaryotic homologue of the eukaryotic protein tubulin -- polymerizes into treadmilling filaments that further organize into a cytoskeletal ring. In vitro, FtsZ filaments can form dynamic chiral assemblies. However, how the active and passive properties of individual filaments relate to these large-scale self-organized structures remains poorly understood. Here, we connect single filament properties with the mesoscopic scale by combining minimal active matter simulations and biochemical reconstitution experiments. We show that density and flexibility of active chiral filaments define their global order. At intermediate densities, curved, flexible filaments organize into chiral rings and polar bands. An effectively nematic organization dominates for high densities and for straight, mutant filaments with increased rigidity. Our predicted phase diagram captures these features quantitatively, demonstrating how the flexibility, density and chirality of active filaments affect their collective behaviour. Our findings shed light on the fundamental properties of active chiral matter and explain how treadmilling FtsZ filaments organize during bacterial cell division. ' acknowledged_ssus: - _id: Bio - _id: LifeSc acknowledgement: 'This work was supported by the European Research Council through grant ERC 2015-StG-679239 and by the Austrian Science Fund (FWF) StandAlone P34607 to M.L., B. P.M. was also supported by the Kanazawa University WPI- NanoLSI Bio-SPM collaborative research program. Z.D. has received funding from Doctoral Programme of the Austrian Academy of Sciences (OeAW): Grant agreement 26360. We thank Jan Brugues (MPI CBG, Dresden, Germany), Andela Saric (ISTA, Klosterneuburg, Austria), Daniel Pearce (Uni Geneva, Switzerland) for valuable scientific input and comments on the manuscript. We are also thankful for the support by the Scientific Service Units (SSU) of IST Austria through resources provided by the Imaging and Optics Facility (IOF) and the Lab Support Facility (LSF). ' article_processing_charge: No author: - first_name: Zuzana full_name: Dunajova, Zuzana id: 4B39F286-F248-11E8-B48F-1D18A9856A87 last_name: Dunajova - first_name: Batirtze full_name: Prats Mateu, Batirtze id: 299FE892-F248-11E8-B48F-1D18A9856A87 last_name: Prats Mateu - first_name: Philipp full_name: Radler, Philipp id: 40136C2A-F248-11E8-B48F-1D18A9856A87 last_name: Radler orcid: '0000-0001-9198-2182 ' - first_name: Keesiang full_name: Lim, Keesiang last_name: Lim - first_name: Dörte full_name: Brandis, Dörte last_name: Brandis - first_name: Philipp full_name: Velicky, Philipp id: 39BDC62C-F248-11E8-B48F-1D18A9856A87 last_name: Velicky orcid: 0000-0002-2340-7431 - first_name: Johann G full_name: Danzl, Johann G id: 42EFD3B6-F248-11E8-B48F-1D18A9856A87 last_name: Danzl orcid: 0000-0001-8559-3973 - first_name: Richard W. full_name: Wong, Richard W. last_name: Wong - first_name: Jens full_name: Elgeti, Jens last_name: Elgeti - first_name: Edouard B full_name: Hannezo, Edouard B id: 3A9DB764-F248-11E8-B48F-1D18A9856A87 last_name: Hannezo orcid: 0000-0001-6005-1561 - first_name: Martin full_name: Loose, Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 citation: ama: Dunajova Z, Prats Mateu B, Radler P, et al. Chiral and nematic phases of flexible active filaments. 2023. doi:10.15479/AT:ISTA:13116 apa: Dunajova, Z., Prats Mateu, B., Radler, P., Lim, K., Brandis, D., Velicky, P., … Loose, M. (2023). Chiral and nematic phases of flexible active filaments. Institute of Science and Technology Austria. https://doi.org/10.15479/AT:ISTA:13116 chicago: Dunajova, Zuzana, Batirtze Prats Mateu, Philipp Radler, Keesiang Lim, Dörte Brandis, Philipp Velicky, Johann G Danzl, et al. “Chiral and Nematic Phases of Flexible Active Filaments.” Institute of Science and Technology Austria, 2023. https://doi.org/10.15479/AT:ISTA:13116. ieee: Z. Dunajova et al., “Chiral and nematic phases of flexible active filaments.” Institute of Science and Technology Austria, 2023. ista: Dunajova Z, Prats Mateu B, Radler P, Lim K, Brandis D, Velicky P, Danzl JG, Wong RW, Elgeti J, Hannezo EB, Loose M. 2023. Chiral and nematic phases of flexible active filaments, Institute of Science and Technology Austria, 10.15479/AT:ISTA:13116. mla: Dunajova, Zuzana, et al. Chiral and Nematic Phases of Flexible Active Filaments. Institute of Science and Technology Austria, 2023, doi:10.15479/AT:ISTA:13116. short: Z. Dunajova, B. Prats Mateu, P. Radler, K. Lim, D. Brandis, P. Velicky, J.G. Danzl, R.W. Wong, J. Elgeti, E.B. Hannezo, M. Loose, (2023). corr_author: '1' date_created: 2023-06-02T12:30:40Z date_published: 2023-07-26T00:00:00Z date_updated: 2026-03-17T12:02:11Z day: '26' ddc: - '539' department: - _id: MaLo - _id: EdHa - _id: JoDa doi: 10.15479/AT:ISTA:13116 ec_funded: 1 file: - access_level: open_access checksum: 4b4ec5d4df7672b3af5ba23b126b171b content_type: application/vnd.openxmlformats-officedocument.wordprocessingml.document creator: pradler date_created: 2023-08-08T11:17:28Z date_updated: 2023-08-08T11:17:28Z file_id: '13983' file_name: ReadMe File.docx file_size: 13111 relation: main_file success: 1 - access_level: open_access checksum: 6f1673d6ae4f547cd49cfe7f87d9ab7e content_type: application/octet-stream creator: pradler date_created: 2023-07-25T06:55:43Z date_updated: 2023-07-25T06:55:43Z file_id: '13298' file_name: TIRF_FtsZ WT 0.625µM.7z.001 file_size: 1499504777 relation: main_file success: 1 - access_level: open_access checksum: bc0dea871af164f7419ac838a34a4162 content_type: application/octet-stream creator: pradler date_created: 2023-07-25T08:31:07Z date_updated: 2023-07-25T08:31:07Z file_id: '13306' file_name: TIRF_FtsZ WT 0.900µM.7z.001 file_size: 3986437211 relation: main_file success: 1 - 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access_level: open_access checksum: 848456b4230c086e8999f57d57dba7f1 content_type: text/csv creator: mloose date_created: 2023-08-03T19:51:54Z date_updated: 2023-08-03T19:51:54Z file_id: '13952' file_name: Fig3a_F200.csv file_size: 59166206 relation: main_file success: 1 file_date_updated: 2023-08-08T11:17:28Z has_accepted_license: '1' month: '07' oa: 1 oa_version: Published Version project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell - _id: fc38323b-9c52-11eb-aca3-ff8afb4a011d grant_number: P34607 name: In vitro reconstitution of bacterial cell division - _id: 34d75525-11ca-11ed-8bc3-89b6307fee9d grant_number: '26360' name: Motile active matter models of migrating cells and chiral filaments publisher: Institute of Science and Technology Austria related_material: record: - id: '13314' relation: used_in_publication status: public - id: '21423' relation: used_in_publication status: public status: public title: Chiral and nematic phases of flexible active filaments tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: research_data user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 year: '2023' ... --- _id: '13314' abstract: - lang: eng text: The emergence of large-scale order in self-organized systems relies on local interactions between individual components. During bacterial cell division, FtsZ—a prokaryotic homologue of the eukaryotic protein tubulin—polymerizes into treadmilling filaments that further organize into a cytoskeletal ring. In vitro, FtsZ filaments can form dynamic chiral assemblies. However, how the active and passive properties of individual filaments relate to these large-scale self-organized structures remains poorly understood. Here we connect single-filament properties with the mesoscopic scale by combining minimal active matter simulations and biochemical reconstitution experiments. We show that the density and flexibility of active chiral filaments define their global order. At intermediate densities, curved, flexible filaments organize into chiral rings and polar bands. An effectively nematic organization dominates for high densities and for straight, mutant filaments with increased rigidity. Our predicted phase diagram quantitatively captures these features, demonstrating how the flexibility, density and chirality of the active filaments affect their collective behaviour. Our findings shed light on the fundamental properties of active chiral matter and explain how treadmilling FtsZ filaments organize during bacterial cell division. acknowledged_ssus: - _id: Bio - _id: LifeSc acknowledgement: 'This work was supported by the European Research Council through grant ERC 2015-StG-679239 and by the Austrian Science Fund (FWF) StandAlone P34607 to M.L., B. P.M. was also supported by the Kanazawa University WPI- NanoLSI Bio-SPM collaborative research program. Z.D. has received funding from Doctoral Programme of the Austrian Academy of Sciences (OeAW): Grant agreement 26360. We thank Jan Brugues (MPI CBG, Dresden, Germany), Andela Saric (ISTA, Klosterneuburg, Austria), Daniel Pearce (Uni Geneva, Switzerland) for valuable scientific input and comments on the manuscript. We are also thankful for the support by the Scientific Service Units (SSU) of IST Austria through resources provided by the Imaging and Optics Facility (IOF) and the Lab Support Facility (LSF).' article_processing_charge: Yes (in subscription journal) article_type: original author: - first_name: Zuzana full_name: Dunajova, Zuzana id: 4B39F286-F248-11E8-B48F-1D18A9856A87 last_name: Dunajova - first_name: Batirtze full_name: Prats Mateu, Batirtze id: 299FE892-F248-11E8-B48F-1D18A9856A87 last_name: Prats Mateu - first_name: Philipp full_name: Radler, Philipp id: 40136C2A-F248-11E8-B48F-1D18A9856A87 last_name: Radler orcid: '0000-0001-9198-2182 ' - first_name: Keesiang full_name: Lim, Keesiang last_name: Lim - first_name: Dörte full_name: Brandis, Dörte id: 21d64d35-f128-11eb-9611-b8bcca7a12fd last_name: Brandis - first_name: Philipp full_name: Velicky, Philipp id: 39BDC62C-F248-11E8-B48F-1D18A9856A87 last_name: Velicky orcid: 0000-0002-2340-7431 - first_name: Johann G full_name: Danzl, Johann G id: 42EFD3B6-F248-11E8-B48F-1D18A9856A87 last_name: Danzl orcid: 0000-0001-8559-3973 - first_name: Richard W. full_name: Wong, Richard W. last_name: Wong - first_name: Jens full_name: Elgeti, Jens last_name: Elgeti - first_name: Edouard B full_name: Hannezo, Edouard B id: 3A9DB764-F248-11E8-B48F-1D18A9856A87 last_name: Hannezo orcid: 0000-0001-6005-1561 - first_name: Martin full_name: Loose, Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 citation: ama: Dunajova Z, Prats Mateu B, Radler P, et al. Chiral and nematic phases of flexible active filaments. Nature Physics. 2023;19:1916-1926. doi:10.1038/s41567-023-02218-w apa: Dunajova, Z., Prats Mateu, B., Radler, P., Lim, K., Brandis, D., Velicky, P., … Loose, M. (2023). Chiral and nematic phases of flexible active filaments. Nature Physics. Springer Nature. https://doi.org/10.1038/s41567-023-02218-w chicago: Dunajova, Zuzana, Batirtze Prats Mateu, Philipp Radler, Keesiang Lim, Dörte Brandis, Philipp Velicky, Johann G Danzl, et al. “Chiral and Nematic Phases of Flexible Active Filaments.” Nature Physics. Springer Nature, 2023. https://doi.org/10.1038/s41567-023-02218-w. ieee: Z. Dunajova et al., “Chiral and nematic phases of flexible active filaments,” Nature Physics, vol. 19. Springer Nature, pp. 1916–1926, 2023. ista: Dunajova Z, Prats Mateu B, Radler P, Lim K, Brandis D, Velicky P, Danzl JG, Wong RW, Elgeti J, Hannezo EB, Loose M. 2023. Chiral and nematic phases of flexible active filaments. Nature Physics. 19, 1916–1926. mla: Dunajova, Zuzana, et al. “Chiral and Nematic Phases of Flexible Active Filaments.” Nature Physics, vol. 19, Springer Nature, 2023, pp. 1916–26, doi:10.1038/s41567-023-02218-w. short: Z. Dunajova, B. Prats Mateu, P. Radler, K. Lim, D. Brandis, P. Velicky, J.G. Danzl, R.W. Wong, J. Elgeti, E.B. Hannezo, M. Loose, Nature Physics 19 (2023) 1916–1926. corr_author: '1' date_created: 2023-07-27T14:44:45Z date_published: 2023-12-01T00:00:00Z date_updated: 2026-03-18T14:11:35Z day: '01' ddc: - '530' department: - _id: JoDa - _id: EdHa - _id: MaLo - _id: GradSch doi: 10.1038/s41567-023-02218-w ec_funded: 1 external_id: isi: - '001178645300041' pmid: - '38075437' file: - access_level: open_access checksum: bc7673ca07d37309013a86166577b2f7 content_type: application/pdf creator: dernst date_created: 2024-01-30T14:28:30Z date_updated: 2024-01-30T14:28:30Z file_id: '14916' file_name: 2023_NaturePhysics_Dunajova.pdf file_size: 22471673 relation: main_file success: 1 file_date_updated: 2024-01-30T14:28:30Z has_accepted_license: '1' intvolume: ' 19' isi: 1 language: - iso: eng month: '12' oa: 1 oa_version: Published Version page: 1916-1926 pmid: 1 project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell - _id: fc38323b-9c52-11eb-aca3-ff8afb4a011d grant_number: P34607 name: In vitro reconstitution of bacterial cell division - _id: 34d75525-11ca-11ed-8bc3-89b6307fee9d grant_number: '26360' name: Motile active matter models of migrating cells and chiral filaments publication: Nature Physics publication_identifier: eissn: - 1745-2481 issn: - 1745-2473 publication_status: published publisher: Springer Nature quality_controlled: '1' related_material: record: - id: '13116' relation: research_data status: public - id: '21423' relation: dissertation_contains status: public - id: '21439' relation: research_data status: public scopus_import: '1' status: public title: Chiral and nematic phases of flexible active filaments tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: 317138e5-6ab7-11ef-aa6d-ffef3953e345 volume: 19 year: '2023' ... --- OA_place: publisher _id: '14280' abstract: - lang: eng text: "Cell division in Escherichia coli is performed by the divisome, a multi-protein complex composed of more than 30 proteins. The divisome spans from the cytoplasm through the inner membrane to the cell wall and the outer membrane. Divisome assembly is initiated by a cytoskeletal structure, the so-called Z-ring, which localizes at the center of the E. coli cell and determines the position of the future cell septum. The Z-ring is composed of the highly conserved bacterial tubulin homologue FtsZ, which forms treadmilling filaments. These filaments are recruited to the inner membrane by FtsA, a highly conserved bacterial actin homologue. FtsA interacts with other proteins in the periplasm and thus connects the cytoplasmic and periplasmic components of the divisome. \r\nA previous model postulated that FtsA regulates maturation of the divisome by switching from an oligomeric, inactive state to a monomeric and active state. This model was based mostly on in vivo studies, as a biochemical characterization of FtsA has been hampered by difficulties in purifying the protein. Here, we studied FtsA using an in vitro reconstitution approach and aimed to answer two questions: (i) How are dynamics from cytoplasmic, treadmilling FtsZ filaments coupled to proteins acting in the periplasmic space and (ii) How does FtsA regulate the maturation of the divisome?\r\nWe found that the cytoplasmic peptides of the transmembrane proteins FtsN and FtsQ interact directly with FtsA and can follow the spatiotemporal signal of FtsA/Z filaments. When we investigated the underlying mechanism by imaging single molecules of FtsNcyto, we found the peptide to interact transiently with FtsA. An in depth analysis of the single molecule trajectories helped to postulate a model where PG synthases follow the dynamics of FtsZ by a diffusion and capture mechanism. \r\nFollowing up on these findings we were interested in how the self-interaction of FtsA changes when it encounters FtsNcyto and if we can confirm the proposed oligomer-monomer switch. For this, we compared the behavior of the previously identified, hyperactive mutant FtsA R286W with wildtype FtsA. The mutant outperforms WT in mirroring and transmitting the spatiotemporal signal of treadmilling FtsZ filaments. Surprisingly however, we found that this was not due to a difference in the self-interaction strength of the two variants, but a difference in their membrane residence time. Furthermore, in contrast to our expectations, upon binding of FtsNcyto the measured self-interaction of FtsA actually increased. \r\nWe propose that FtsNcyto induces a rearrangement of the oligomeric architecture of FtsA. In further consequence this change leads to more persistent FtsZ filaments which results in a defined signalling zone, allowing formation of the mature divisome. The observed difference between FtsA WT and R286W is due to the vastly different membrane turnover of the proteins. R286W cycles 5-10x faster compared to WT which allows to sample FtsZ filaments at faster frequencies. These findings can explain the observed differences in toxicity for overexpression of FtsA WT and R286W and help to understand how FtsA regulates divisome maturation." acknowledged_ssus: - _id: Bio - _id: LifeSc alternative_title: - ISTA Thesis article_processing_charge: No author: - first_name: Philipp full_name: Radler, Philipp id: 40136C2A-F248-11E8-B48F-1D18A9856A87 last_name: Radler orcid: '0000-0001-9198-2182 ' citation: ama: Radler P. Spatiotemporal signaling during assembly of the bacterial divisome. 2023. doi:10.15479/at:ista:14280 apa: Radler, P. (2023). Spatiotemporal signaling during assembly of the bacterial divisome. Institute of Science and Technology Austria. https://doi.org/10.15479/at:ista:14280 chicago: Radler, Philipp. “Spatiotemporal Signaling during Assembly of the Bacterial Divisome.” Institute of Science and Technology Austria, 2023. https://doi.org/10.15479/at:ista:14280. ieee: P. Radler, “Spatiotemporal signaling during assembly of the bacterial divisome,” Institute of Science and Technology Austria, 2023. ista: Radler P. 2023. Spatiotemporal signaling during assembly of the bacterial divisome. Institute of Science and Technology Austria. mla: Radler, Philipp. Spatiotemporal Signaling during Assembly of the Bacterial Divisome. Institute of Science and Technology Austria, 2023, doi:10.15479/at:ista:14280. short: P. Radler, Spatiotemporal Signaling during Assembly of the Bacterial Divisome, Institute of Science and Technology Austria, 2023. corr_author: '1' date_created: 2023-09-06T10:58:25Z date_published: 2023-09-25T00:00:00Z date_updated: 2026-04-07T14:06:05Z day: '25' ddc: - '572' degree_awarded: PhD department: - _id: GradSch - _id: MaLo doi: 10.15479/at:ista:14280 ec_funded: 1 file: - access_level: closed checksum: 87eef11fbc5c7df0826f12a3a629b444 content_type: application/vnd.openxmlformats-officedocument.wordprocessingml.document creator: pradler date_created: 2023-10-04T10:11:53Z date_updated: 2024-10-05T22:30:03Z embargo_to: open_access file_id: '14390' file_name: PhD Thesis_Philipp Radler_20231004.docx file_size: 114932847 relation: source_file - access_level: open_access checksum: 3253e099b7126469d941fd9419d68b4f content_type: application/pdf creator: pradler date_created: 2023-10-04T10:11:21Z date_updated: 2024-10-05T22:30:03Z embargo: 2024-10-04 file_id: '14391' file_name: PhD Thesis_Philipp Radler_20231004.pdf file_size: 37838778 relation: main_file file_date_updated: 2024-10-05T22:30:03Z has_accepted_license: '1' keyword: - Cell Division - Reconstitution - FtsZ - FtsA - Divisome - E.coli language: - iso: eng month: '09' oa: 1 oa_version: Published Version page: '156' project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell - _id: fc38323b-9c52-11eb-aca3-ff8afb4a011d grant_number: P34607 name: In vitro reconstitution of bacterial cell division - _id: 2596EAB6-B435-11E9-9278-68D0E5697425 grant_number: ALTF 2015-1163 name: Synthesis of bacterial cell wall - _id: 259B655A-B435-11E9-9278-68D0E5697425 grant_number: LT000824/2016 name: Reconstitution of bacterial cell wall synthesis publication_identifier: isbn: - 978-3-99078-033-6 issn: - 2663-337X publication_status: published publisher: Institute of Science and Technology Austria related_material: record: - id: '10934' relation: research_data status: public - id: '11373' relation: part_of_dissertation status: public - id: '7387' relation: part_of_dissertation status: public status: public supervisor: - first_name: Martin full_name: Loose, Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 title: Spatiotemporal signaling during assembly of the bacterial divisome tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: dissertation user_id: ba8df636-2132-11f1-aed0-ed93e2281fdd year: '2023' ... --- _id: '10934' abstract: - lang: eng text: 'FtsA is crucial for assembly of the E. coli divisome, as it dynamically links cytoplasmic FtsZ filaments with transmembrane cell division proteins. FtsA allegedly initiates cell division by switching from an inactive polymeric to an active monomeric confirmation, which recruits downstream proteins and stabilizes FtsZ filaments. Here, we use biochemical reconstitution experiments combined with quantitative fluorescence microscopy to study divisome activation in vitro. We compare wildtype-FtsA with FtsA-R286W, a constantly active gain-of-function mutant and find that R286W outperforms the wildtype protein in replicating FtsZ treadmilling dynamics, stabilizing FtsZ filaments and recruiting FtsN. We attribute these differences to a faster membrane exchange of FtsA-R286W and its higher packing density below FtsZ filaments. Using FRET microscopy, we find that FtsN binding does not compete with, but promotes FtsA self-interaction. Our findings suggest a model where FtsA always forms dynamic polymers on the membrane, which re-organize during assembly and activation of the divisome. ' acknowledged_ssus: - _id: Bio - _id: LifeSc acknowledgement: We acknowledge members of the Loose laboratory at IST Austria for helpful discussions—in particular L. Lindorfer for his assistance with cloning and purifications. We thank J. Löwe and T. Nierhaus (MRC-LMB Cambridge, UK) for sharing unpublished work and helpful discussions, as well as D. Vavylonis and D. Rutkowski (Lehigh University, Bethlehem, PA, USA) as well as S. Martin (University of Lausanne, Switzerland) for sharing their code for FRAP analysis. We are also thankful for the support by the Scientific Service Units (SSU) of IST Austria through resources provided by the Imaging and Optics Facility (IOF) and the Lab Support Facility (LSF). This work was supported by the European Research Council through grant ERC 2015-StG-679239 and by the Austrian Science Fund (FWF) StandAlone P34607 to M.L. and HFSP LT 000824/2016-L4 to N.B. For the purpose of open access, we have applied a CC BY public copyright licence to any Author Accepted Manuscript version arising from this submission. article_processing_charge: No author: - first_name: Philipp full_name: Radler, Philipp id: 40136C2A-F248-11E8-B48F-1D18A9856A87 last_name: Radler orcid: ' 0000-0001-9198-2182 ' citation: ama: Radler P. In vitro reconstitution of Escherichia coli divisome activation. 2022. doi:10.15479/AT:ISTA:10934 apa: Radler, P. (2022). In vitro reconstitution of Escherichia coli divisome activation. Institute of Science and Technology Austria. https://doi.org/10.15479/AT:ISTA:10934 chicago: Radler, Philipp. “In Vitro Reconstitution of Escherichia Coli Divisome Activation.” Institute of Science and Technology Austria, 2022. https://doi.org/10.15479/AT:ISTA:10934. ieee: P. Radler, “In vitro reconstitution of Escherichia coli divisome activation.” Institute of Science and Technology Austria, 2022. ista: Radler P. 2022. In vitro reconstitution of Escherichia coli divisome activation, Institute of Science and Technology Austria, 10.15479/AT:ISTA:10934. mla: Radler, Philipp. In Vitro Reconstitution of Escherichia Coli Divisome Activation. Institute of Science and Technology Austria, 2022, doi:10.15479/AT:ISTA:10934. short: P. Radler, (2022). contributor: - contributor_type: supervisor first_name: Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 - contributor_type: researcher first_name: Christoph M id: 4DF26D8C-F248-11E8-B48F-1D18A9856A87 last_name: Sommer - contributor_type: researcher first_name: Paulo last_name: Caldas - contributor_type: researcher first_name: David id: B9577E20-AA38-11E9-AC9A-0930E6697425 last_name: Michalik - contributor_type: researcher first_name: Natalia last_name: Baranova corr_author: '1' date_created: 2022-03-31T11:32:32Z date_published: 2022-04-05T00:00:00Z date_updated: 2026-04-28T22:30:27Z day: '05' ddc: - '572' department: - _id: GradSch - _id: MaLo doi: 10.15479/AT:ISTA:10934 ec_funded: 1 file: - access_level: open_access checksum: 52d50202e04e9daa618a58e686d8ab58 content_type: application/vnd.openxmlformats-officedocument.wordprocessingml.document creator: pradler date_created: 2022-04-22T10:15:19Z date_updated: 2022-04-22T10:15:19Z file_id: '11328' file_name: Inventory for Data repository.docx file_size: 13469 relation: main_file success: 1 - 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access_level: open_access checksum: 3f4928a36e1b1f295054668060df3079 content_type: application/octet-stream creator: pradler date_created: 2022-04-05T08:51:55Z date_updated: 2022-04-05T08:51:55Z file_id: '10953' file_name: Raw Microscopy_FRET FtsA His6 + FtsN & FtsZ_01.z01 file_size: 4294960000 relation: main_file success: 1 file_date_updated: 2022-04-22T10:15:19Z has_accepted_license: '1' keyword: - Bacterial cell division - in vitro reconstitution - FtsZ - FtsN - FtsA month: '04' oa: 1 oa_version: Submitted Version project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell - _id: fc38323b-9c52-11eb-aca3-ff8afb4a011d grant_number: P34607 name: In vitro reconstitution of bacterial cell division publisher: Institute of Science and Technology Austria related_material: link: - description: A custom written code (FRAPdiff) to quantify the Off binding rate and Diffusion coefficient of membrane bound proteins. Written by Christoph Sommer. relation: software url: https://doi.org/10.5281/zenodo.6400639 record: - id: '11373' relation: used_in_publication status: public - id: '14280' relation: used_in_publication status: public status: public title: In vitro reconstitution of Escherichia coli divisome activation tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: research_data user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 year: '2022' ... --- _id: '11373' abstract: - lang: eng text: The actin-homologue FtsA is essential for E. coli cell division, as it links FtsZ filaments in the Z-ring to transmembrane proteins. FtsA is thought to initiate cell constriction by switching from an inactive polymeric to an active monomeric conformation, which recruits downstream proteins and stabilizes the Z-ring. However, direct biochemical evidence for this mechanism is missing. Here, we use reconstitution experiments and quantitative fluorescence microscopy to study divisome activation in vitro. By comparing wild-type FtsA with FtsA R286W, we find that this hyperactive mutant outperforms FtsA WT in replicating FtsZ treadmilling dynamics, FtsZ filament stabilization and recruitment of FtsN. We could attribute these differences to a faster exchange and denser packing of FtsA R286W below FtsZ filaments. Using FRET microscopy, we also find that FtsN binding promotes FtsA self-interaction. We propose that in the active divisome FtsA and FtsN exist as a dynamic copolymer that follows treadmilling filaments of FtsZ. acknowledged_ssus: - _id: Bio - _id: LifeSc acknowledgement: We acknowledge members of the Loose laboratory at IST Austria for helpful discussions—in particular L. Lindorfer for his assistance with cloning and purifications. We thank J. Löwe and T. Nierhaus (MRC-LMB Cambridge, UK) for sharing unpublished work and helpful discussions, as well as D. Vavylonis and D. Rutkowski (Lehigh University, Bethlehem, PA, USA) and S. Martin (University of Lausanne, Switzerland) for sharing their code for FRAP analysis. We are also thankful for the support by the Scientific Service Units (SSU) of IST Austria through resources provided by the Imaging and Optics Facility (IOF) and the Lab Support Facility (LSF). This work was supported by the European Research Council through grant ERC 2015-StG-679239 and by the Austrian Science Fund (FWF) StandAlone P34607 to M.L. and HFSP LT 000824/2016-L4 to N.B. For the purpose of open access, we have applied a CC BY public copyright licence to any Author Accepted Manuscript version arising from this submission. article_number: '2635' article_processing_charge: No article_type: original author: - first_name: Philipp full_name: Radler, Philipp id: 40136C2A-F248-11E8-B48F-1D18A9856A87 last_name: Radler orcid: '0000-0001-9198-2182 ' - first_name: Natalia S. full_name: Baranova, Natalia S. id: 38661662-F248-11E8-B48F-1D18A9856A87 last_name: Baranova orcid: 0000-0002-3086-9124 - first_name: Paulo R full_name: Dos Santos Caldas, Paulo R id: 38FCDB4C-F248-11E8-B48F-1D18A9856A87 last_name: Dos Santos Caldas orcid: 0000-0001-6730-4461 - first_name: Christoph M full_name: Sommer, Christoph M id: 4DF26D8C-F248-11E8-B48F-1D18A9856A87 last_name: Sommer orcid: 0000-0003-1216-9105 - first_name: Maria D full_name: Lopez Pelegrin, Maria D id: 319AA9CE-F248-11E8-B48F-1D18A9856A87 last_name: Lopez Pelegrin - first_name: David full_name: Michalik, David id: B9577E20-AA38-11E9-AC9A-0930E6697425 last_name: Michalik - first_name: Martin full_name: Loose, Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 citation: ama: Radler P, Baranova NS, Dos Santos Caldas PR, et al. In vitro reconstitution of Escherichia coli divisome activation. Nature Communications. 2022;13. doi:10.1038/s41467-022-30301-y apa: Radler, P., Baranova, N. S., Dos Santos Caldas, P. R., Sommer, C. M., Lopez Pelegrin, M. D., Michalik, D., & Loose, M. (2022). In vitro reconstitution of Escherichia coli divisome activation. Nature Communications. Springer Nature. https://doi.org/10.1038/s41467-022-30301-y chicago: Radler, Philipp, Natalia S. Baranova, Paulo R Dos Santos Caldas, Christoph M Sommer, Maria D Lopez Pelegrin, David Michalik, and Martin Loose. “In Vitro Reconstitution of Escherichia Coli Divisome Activation.” Nature Communications. Springer Nature, 2022. https://doi.org/10.1038/s41467-022-30301-y. ieee: P. Radler et al., “In vitro reconstitution of Escherichia coli divisome activation,” Nature Communications, vol. 13. Springer Nature, 2022. ista: Radler P, Baranova NS, Dos Santos Caldas PR, Sommer CM, Lopez Pelegrin MD, Michalik D, Loose M. 2022. In vitro reconstitution of Escherichia coli divisome activation. Nature Communications. 13, 2635. mla: Radler, Philipp, et al. “In Vitro Reconstitution of Escherichia Coli Divisome Activation.” Nature Communications, vol. 13, 2635, Springer Nature, 2022, doi:10.1038/s41467-022-30301-y. short: P. Radler, N.S. Baranova, P.R. Dos Santos Caldas, C.M. Sommer, M.D. Lopez Pelegrin, D. Michalik, M. Loose, Nature Communications 13 (2022). corr_author: '1' date_created: 2022-05-13T09:06:28Z date_published: 2022-05-12T00:00:00Z date_updated: 2026-04-28T22:30:27Z day: '12' ddc: - '570' department: - _id: MaLo doi: 10.1038/s41467-022-30301-y ec_funded: 1 external_id: isi: - '000795171100037' file: - access_level: open_access checksum: 5af863ee1b95a0710f6ee864d68dc7a6 content_type: application/pdf creator: dernst date_created: 2022-05-13T09:10:51Z date_updated: 2022-05-13T09:10:51Z file_id: '11374' file_name: 2022_NatureCommunications_Radler.pdf file_size: 6945191 relation: main_file success: 1 file_date_updated: 2022-05-13T09:10:51Z has_accepted_license: '1' intvolume: ' 13' isi: 1 keyword: - General Physics and Astronomy - General Biochemistry - Genetics and Molecular Biology - General Chemistry language: - iso: eng month: '05' oa: 1 oa_version: Published Version project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell - _id: fc38323b-9c52-11eb-aca3-ff8afb4a011d grant_number: P34607 name: In vitro reconstitution of bacterial cell division publication: Nature Communications publication_identifier: issn: - 2041-1723 publication_status: published publisher: Springer Nature quality_controlled: '1' related_material: link: - relation: erratum url: https://doi.org/10.1038/s41467-022-34485-1 record: - id: '10934' relation: research_data status: public - id: '14280' relation: dissertation_contains status: public scopus_import: '1' status: public title: In vitro reconstitution of Escherichia coli divisome activation tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: 4359f0d1-fa6c-11eb-b949-802e58b17ae8 volume: 13 year: '2022' ... --- _id: '9243' abstract: - lang: eng text: Peptidoglycan is an essential component of the bacterial cell envelope that surrounds the cytoplasmic membrane to protect the cell from osmotic lysis. Important antibiotics such as β-lactams and glycopeptides target peptidoglycan biosynthesis. Class A penicillin-binding proteins (PBPs) are bifunctional membrane-bound peptidoglycan synthases that polymerize glycan chains and connect adjacent stem peptides by transpeptidation. How these enzymes work in their physiological membrane environment is poorly understood. Here, we developed a novel Förster resonance energy transfer-based assay to follow in real time both reactions of class A PBPs reconstituted in liposomes or supported lipid bilayers and applied this assay with PBP1B homologues from Escherichia coli, Pseudomonas aeruginosa, and Acinetobacter baumannii in the presence or absence of their cognate lipoprotein activator. Our assay will allow unravelling the mechanisms of peptidoglycan synthesis in a lipid-bilayer environment and can be further developed to be used for high-throughput screening for new antimicrobials. acknowledgement: 'We thank Alexander Egan (Newcastle University) for purified proteins LpoB(sol) and LpoPPa(sol), Federico Corona (Newcastle University) for purified MepM, and Oliver Birkholz and Jacob Piehler (Department of Biology and Center of Cellular Nanoanalytics, University of Osnabru¨ ck) for their help with PBP1B reconstitution into polymer-SLBs and initial guidance on single particle tracking. We also acknowledge Christian P Richter and Changjiang You (Department of Biology and Center of Cellular Nanoanalytics, University of Osnabru¨ ck) for providing SLIMfast software and tris-DODA-NTA reagent, respectively. This work was funded by the BBSRC grant BB/R017409/1 (to WV), the European Research Council through grant ERC-2015-StG-679239 (to ML), and long-term fellowships HFSP LT 000824/2016-L4 and EMBO ALTF 1163–2015 (to NB). ' article_number: 1-32 article_processing_charge: No article_type: original author: - first_name: Víctor M. full_name: Hernández-Rocamora, Víctor M. last_name: Hernández-Rocamora - first_name: Natalia S. full_name: Baranova, Natalia S. id: 38661662-F248-11E8-B48F-1D18A9856A87 last_name: Baranova orcid: 0000-0002-3086-9124 - first_name: Katharina full_name: Peters, Katharina last_name: Peters - first_name: Eefjan full_name: Breukink, Eefjan last_name: Breukink - first_name: Martin full_name: Loose, Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 - first_name: Waldemar full_name: Vollmer, Waldemar last_name: Vollmer citation: ama: Hernández-Rocamora VM, Baranova NS, Peters K, Breukink E, Loose M, Vollmer W. Real time monitoring of peptidoglycan synthesis by membrane-reconstituted penicillin binding proteins. eLife. 2021;10. doi:10.7554/eLife.61525 apa: Hernández-Rocamora, V. M., Baranova, N. S., Peters, K., Breukink, E., Loose, M., & Vollmer, W. (2021). Real time monitoring of peptidoglycan synthesis by membrane-reconstituted penicillin binding proteins. ELife. eLife Sciences Publications. https://doi.org/10.7554/eLife.61525 chicago: Hernández-Rocamora, Víctor M., Natalia S. Baranova, Katharina Peters, Eefjan Breukink, Martin Loose, and Waldemar Vollmer. “Real Time Monitoring of Peptidoglycan Synthesis by Membrane-Reconstituted Penicillin Binding Proteins.” ELife. eLife Sciences Publications, 2021. https://doi.org/10.7554/eLife.61525. ieee: V. M. Hernández-Rocamora, N. S. Baranova, K. Peters, E. Breukink, M. Loose, and W. Vollmer, “Real time monitoring of peptidoglycan synthesis by membrane-reconstituted penicillin binding proteins,” eLife, vol. 10. eLife Sciences Publications, 2021. ista: Hernández-Rocamora VM, Baranova NS, Peters K, Breukink E, Loose M, Vollmer W. 2021. Real time monitoring of peptidoglycan synthesis by membrane-reconstituted penicillin binding proteins. eLife. 10, 1–32. mla: Hernández-Rocamora, Víctor M., et al. “Real Time Monitoring of Peptidoglycan Synthesis by Membrane-Reconstituted Penicillin Binding Proteins.” ELife, vol. 10, 1–32, eLife Sciences Publications, 2021, doi:10.7554/eLife.61525. short: V.M. Hernández-Rocamora, N.S. Baranova, K. Peters, E. Breukink, M. Loose, W. Vollmer, ELife 10 (2021). date_created: 2021-03-14T23:01:33Z date_published: 2021-02-24T00:00:00Z date_updated: 2024-10-22T10:04:21Z day: '24' ddc: - '570' department: - _id: MaLo doi: 10.7554/eLife.61525 ec_funded: 1 external_id: isi: - '000627596400001' file: - access_level: open_access checksum: 79897a09bfecd9914d39c4aea2841855 content_type: application/pdf creator: dernst date_created: 2021-03-22T07:36:08Z date_updated: 2021-03-22T07:36:08Z file_id: '9268' file_name: 2021_eLife_HernandezRocamora.pdf file_size: 2314698 relation: main_file success: 1 file_date_updated: 2021-03-22T07:36:08Z has_accepted_license: '1' intvolume: ' 10' isi: 1 language: - iso: eng month: '02' oa: 1 oa_version: Published Version project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell - _id: 2596EAB6-B435-11E9-9278-68D0E5697425 grant_number: ALTF 2015-1163 name: Synthesis of bacterial cell wall - _id: 259B655A-B435-11E9-9278-68D0E5697425 grant_number: LT000824/2016 name: Reconstitution of bacterial cell wall synthesis publication: eLife publication_identifier: eissn: - 2050-084X publication_status: published publisher: eLife Sciences Publications quality_controlled: '1' scopus_import: '1' status: public title: Real time monitoring of peptidoglycan synthesis by membrane-reconstituted penicillin binding proteins tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: 4359f0d1-fa6c-11eb-b949-802e58b17ae8 volume: 10 year: '2021' ... --- _id: '9414' abstract: - lang: eng text: Microtubule plus-end depolymerization rate is a potentially important target of physiological regulation, but it has been challenging to measure, so its role in spatial organization is poorly understood. Here we apply a method for tracking plus ends based on time difference imaging to measure depolymerization rates in large interphase asters growing in Xenopus egg extract. We observed strong spatial regulation of depolymerization rates, which were higher in the aster interior compared with the periphery, and much less regulation of polymerization or catastrophe rates. We interpret these data in terms of a limiting component model, where aster growth results in lower levels of soluble tubulin and microtubule-associated proteins (MAPs) in the interior cytosol compared with that at the periphery. The steady-state polymer fraction of tubulin was ∼30%, so tubulin is not strongly depleted in the aster interior. We propose that the limiting component for microtubule assembly is a MAP that inhibits depolymerization, and that egg asters are tuned to low microtubule density. acknowledgement: The authors thank the members of Mitchison, Brugués, and Jay Gatlin groups (University of Wyoming) for discussions. We thank Heino Andreas (MPI-CBG) for frog maintenance. We thank Nikon for microscopy support at Marine Biological Laboratory (MBL). K.I. was supported by fellowships from the Honjo International Scholarship Foundation and Center of Systems Biology Dresden. F.D. was supported by the DIGGS-BB fellowship provided by the German Research Foundation (DFG). P.C. is supported by a Boehringer Ingelheim Fonds PhD fellowship. J.F.P. was supported by a fellowship from the Fannie and John Hertz Foundation. M.L.’s research is supported by European Research Council (ERC) Grant no. ERC-2015-StG-679239. J.B.’s research is supported by the Human Frontiers Science Program (CDA00074/2014). T.J.M.’s research is supported by National Institutes of Health Grant no. R35GM131753. article_processing_charge: No article_type: original author: - first_name: Keisuke full_name: Ishihara, Keisuke last_name: Ishihara - first_name: Franziska full_name: Decker, Franziska last_name: Decker - first_name: Paulo R full_name: Dos Santos Caldas, Paulo R id: 38FCDB4C-F248-11E8-B48F-1D18A9856A87 last_name: Dos Santos Caldas orcid: 0000-0001-6730-4461 - first_name: James F. full_name: Pelletier, James F. last_name: Pelletier - first_name: Martin full_name: Loose, Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 - first_name: Jan full_name: Brugués, Jan last_name: Brugués - first_name: Timothy J. full_name: Mitchison, Timothy J. last_name: Mitchison citation: ama: Ishihara K, Decker F, Dos Santos Caldas PR, et al. Spatial variation of microtubule depolymerization in large asters. Molecular Biology of the Cell. 2021;32(9):869-879. doi:10.1091/MBC.E20-11-0723 apa: Ishihara, K., Decker, F., Dos Santos Caldas, P. R., Pelletier, J. F., Loose, M., Brugués, J., & Mitchison, T. J. (2021). Spatial variation of microtubule depolymerization in large asters. Molecular Biology of the Cell. American Society for Cell Biology. https://doi.org/10.1091/MBC.E20-11-0723 chicago: Ishihara, Keisuke, Franziska Decker, Paulo R Dos Santos Caldas, James F. Pelletier, Martin Loose, Jan Brugués, and Timothy J. Mitchison. “Spatial Variation of Microtubule Depolymerization in Large Asters.” Molecular Biology of the Cell. American Society for Cell Biology, 2021. https://doi.org/10.1091/MBC.E20-11-0723. ieee: K. Ishihara et al., “Spatial variation of microtubule depolymerization in large asters,” Molecular Biology of the Cell, vol. 32, no. 9. American Society for Cell Biology, pp. 869–879, 2021. ista: Ishihara K, Decker F, Dos Santos Caldas PR, Pelletier JF, Loose M, Brugués J, Mitchison TJ. 2021. Spatial variation of microtubule depolymerization in large asters. Molecular Biology of the Cell. 32(9), 869–879. mla: Ishihara, Keisuke, et al. “Spatial Variation of Microtubule Depolymerization in Large Asters.” Molecular Biology of the Cell, vol. 32, no. 9, American Society for Cell Biology, 2021, pp. 869–79, doi:10.1091/MBC.E20-11-0723. short: K. Ishihara, F. Decker, P.R. Dos Santos Caldas, J.F. Pelletier, M. Loose, J. Brugués, T.J. Mitchison, Molecular Biology of the Cell 32 (2021) 869–879. date_created: 2021-05-23T22:01:45Z date_published: 2021-04-19T00:00:00Z date_updated: 2025-04-14T07:21:30Z day: '19' department: - _id: MaLo doi: 10.1091/MBC.E20-11-0723 ec_funded: 1 external_id: isi: - '000641574700005' pmid: - '33439671' intvolume: ' 32' isi: 1 issue: '9' language: - iso: eng main_file_link: - open_access: '1' url: https://www.molbiolcell.org/doi/10.1091/mbc.E20-11-0723 month: '04' oa: 1 oa_version: Published Version page: 869-879 pmid: 1 project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell - _id: 260D98C8-B435-11E9-9278-68D0E5697425 name: Reconstitution of Bacterial Cell Division Using Purified Components publication: Molecular Biology of the Cell publication_identifier: eissn: - 1939-4586 issn: - 1059-1524 publication_status: published publisher: American Society for Cell Biology quality_controlled: '1' scopus_import: '1' status: public title: Spatial variation of microtubule depolymerization in large asters tmp: image: /images/cc_by_nc_sa.png legal_code_url: https://creativecommons.org/licenses/by-nc-sa/3.0/legalcode name: Creative Commons Attribution-NonCommercial-ShareAlike 3.0 Unported (CC BY-NC-SA 3.0) short: CC BY-NC-SA (3.0) type: journal_article user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 volume: 32 year: '2021' ... --- _id: '9907' abstract: - lang: eng text: 'DivIVA is a protein initially identified as a spatial regulator of cell division in the model organism Bacillus subtilis, but its homologues are present in many other Gram-positive bacteria, including Clostridia species. Besides its role as topological regulator of the Min system during bacterial cell division, DivIVA is involved in chromosome segregation during sporulation, genetic competence, and cell wall synthesis. DivIVA localizes to regions of high membrane curvature, such as the cell poles and cell division site, where it recruits distinct binding partners. Previously, it was suggested that negative curvature sensing is the main mechanism by which DivIVA binds to these specific regions. Here, we show that Clostridioides difficile DivIVA binds preferably to membranes containing negatively charged phospholipids, especially cardiolipin. Strikingly, we observed that upon binding, DivIVA modifies the lipid distribution and induces changes to lipid bilayers containing cardiolipin. Our observations indicate that DivIVA might play a more complex and so far unknown active role during the formation of the cell division septal membrane. ' acknowledged_ssus: - _id: Bio - _id: LifeSc acknowledgement: "We thank Daniela Krajˇcíkova, Katarína Muchová, Zuzana Chromíkova and other members of Barák’s laboratory for useful discussions, suggestions and help. Special thanks also to Emília Chovancová for technical support. We are grateful to Juraj Labaj for drawing the model and for help with graphics. Many thanks to all members of Loose’s laboratory: Maria del Mar\r\nLópez, Paulo Caldas, Philipp Radler, and other members of the Loose’s laboratory for sharing their knowledge of SLB preparation and TIRF experiment chambers, for sharing coverslips and for help with the TIRF microscope and data analysis. We also thank the members of the Dept. of Biochemistry of Biomembranes at the Institute of Animal Biochemistry and Genetics, CBs SAS for their help with preparing the lipid mixtures. We thank J. Bauer for critically reading the manuscript." article_number: '8350' article_processing_charge: Yes article_type: original author: - first_name: Naďa full_name: Labajová, Naďa last_name: Labajová - first_name: Natalia S. full_name: Baranova, Natalia S. id: 38661662-F248-11E8-B48F-1D18A9856A87 last_name: Baranova orcid: 0000-0002-3086-9124 - first_name: Miroslav full_name: Jurásek, Miroslav last_name: Jurásek - first_name: Robert full_name: Vácha, Robert last_name: Vácha - first_name: Martin full_name: Loose, Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 - first_name: Imrich full_name: Barák, Imrich last_name: Barák citation: ama: Labajová N, Baranova NS, Jurásek M, Vácha R, Loose M, Barák I. Cardiolipin-containing lipid membranes attract the bacterial cell division protein diviva. International Journal of Molecular Sciences. 2021;22(15). doi:10.3390/ijms22158350 apa: Labajová, N., Baranova, N. S., Jurásek, M., Vácha, R., Loose, M., & Barák, I. (2021). Cardiolipin-containing lipid membranes attract the bacterial cell division protein diviva. International Journal of Molecular Sciences. MDPI. https://doi.org/10.3390/ijms22158350 chicago: Labajová, Naďa, Natalia S. Baranova, Miroslav Jurásek, Robert Vácha, Martin Loose, and Imrich Barák. “Cardiolipin-Containing Lipid Membranes Attract the Bacterial Cell Division Protein Diviva.” International Journal of Molecular Sciences. MDPI, 2021. https://doi.org/10.3390/ijms22158350. ieee: N. Labajová, N. S. Baranova, M. Jurásek, R. Vácha, M. Loose, and I. Barák, “Cardiolipin-containing lipid membranes attract the bacterial cell division protein diviva,” International Journal of Molecular Sciences, vol. 22, no. 15. MDPI, 2021. ista: Labajová N, Baranova NS, Jurásek M, Vácha R, Loose M, Barák I. 2021. Cardiolipin-containing lipid membranes attract the bacterial cell division protein diviva. International Journal of Molecular Sciences. 22(15), 8350. mla: Labajová, Naďa, et al. “Cardiolipin-Containing Lipid Membranes Attract the Bacterial Cell Division Protein Diviva.” International Journal of Molecular Sciences, vol. 22, no. 15, 8350, MDPI, 2021, doi:10.3390/ijms22158350. short: N. Labajová, N.S. Baranova, M. Jurásek, R. Vácha, M. Loose, I. Barák, International Journal of Molecular Sciences 22 (2021). date_created: 2021-08-15T22:01:27Z date_published: 2021-08-01T00:00:00Z date_updated: 2025-07-10T12:02:05Z day: '01' ddc: - '570' department: - _id: MaLo doi: 10.3390/ijms22158350 ec_funded: 1 external_id: isi: - '000681815400001' pmid: - '34361115' file: - access_level: open_access checksum: a4bc06e9a2c803ceff5a91f10b174054 content_type: application/pdf creator: asandaue date_created: 2021-08-16T09:35:56Z date_updated: 2021-08-16T09:35:56Z file_id: '9923' file_name: 2021_InternationalJournalOfMolecularSciences_Labajová .pdf file_size: 6132410 relation: main_file success: 1 file_date_updated: 2021-08-16T09:35:56Z has_accepted_license: '1' intvolume: ' 22' isi: 1 issue: '15' language: - iso: eng month: '08' oa: 1 oa_version: Published Version pmid: 1 project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell publication: International Journal of Molecular Sciences publication_identifier: eissn: - 1422-0067 issn: - 1661-6596 publication_status: published publisher: MDPI quality_controlled: '1' scopus_import: '1' status: public title: Cardiolipin-containing lipid membranes attract the bacterial cell division protein diviva tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 volume: 22 year: '2021' ... --- _id: '7572' abstract: - lang: eng text: The polymerization–depolymerization dynamics of cytoskeletal proteins play essential roles in the self-organization of cytoskeletal structures, in eukaryotic as well as prokaryotic cells. While advances in fluorescence microscopy and in vitro reconstitution experiments have helped to study the dynamic properties of these complex systems, methods that allow to collect and analyze large quantitative datasets of the underlying polymer dynamics are still missing. Here, we present a novel image analysis workflow to study polymerization dynamics of active filaments in a nonbiased, highly automated manner. Using treadmilling filaments of the bacterial tubulin FtsZ as an example, we demonstrate that our method is able to specifically detect, track and analyze growth and shrinkage of polymers, even in dense networks of filaments. We believe that this automated method can facilitate the analysis of a large variety of dynamic cytoskeletal systems, using standard time-lapse movies obtained from experiments in vitro as well as in the living cell. Moreover, we provide scripts implementing this method as supplementary material. alternative_title: - Methods in Cell Biology article_processing_charge: No author: - first_name: Paulo R full_name: Dos Santos Caldas, Paulo R id: 38FCDB4C-F248-11E8-B48F-1D18A9856A87 last_name: Dos Santos Caldas orcid: 0000-0001-6730-4461 - first_name: Philipp full_name: Radler, Philipp id: 40136C2A-F248-11E8-B48F-1D18A9856A87 last_name: Radler orcid: '0000-0001-9198-2182 ' - first_name: Christoph M full_name: Sommer, Christoph M id: 4DF26D8C-F248-11E8-B48F-1D18A9856A87 last_name: Sommer orcid: 0000-0003-1216-9105 - first_name: Martin full_name: Loose, Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 citation: ama: 'Dos Santos Caldas PR, Radler P, Sommer CM, Loose M. Computational analysis of filament polymerization dynamics in cytoskeletal networks. In: Tran P, ed. Methods in Cell Biology. Vol 158. Elsevier; 2020:145-161. doi:10.1016/bs.mcb.2020.01.006' apa: Dos Santos Caldas, P. R., Radler, P., Sommer, C. M., & Loose, M. (2020). Computational analysis of filament polymerization dynamics in cytoskeletal networks. In P. Tran (Ed.), Methods in Cell Biology (Vol. 158, pp. 145–161). Elsevier. https://doi.org/10.1016/bs.mcb.2020.01.006 chicago: Dos Santos Caldas, Paulo R, Philipp Radler, Christoph M Sommer, and Martin Loose. “Computational Analysis of Filament Polymerization Dynamics in Cytoskeletal Networks.” In Methods in Cell Biology, edited by Phong Tran, 158:145–61. Elsevier, 2020. https://doi.org/10.1016/bs.mcb.2020.01.006. ieee: P. R. Dos Santos Caldas, P. Radler, C. M. Sommer, and M. Loose, “Computational analysis of filament polymerization dynamics in cytoskeletal networks,” in Methods in Cell Biology, vol. 158, P. Tran, Ed. Elsevier, 2020, pp. 145–161. ista: 'Dos Santos Caldas PR, Radler P, Sommer CM, Loose M. 2020.Computational analysis of filament polymerization dynamics in cytoskeletal networks. In: Methods in Cell Biology. Methods in Cell Biology, vol. 158, 145–161.' mla: Dos Santos Caldas, Paulo R., et al. “Computational Analysis of Filament Polymerization Dynamics in Cytoskeletal Networks.” Methods in Cell Biology, edited by Phong Tran, vol. 158, Elsevier, 2020, pp. 145–61, doi:10.1016/bs.mcb.2020.01.006. short: P.R. Dos Santos Caldas, P. Radler, C.M. Sommer, M. Loose, in:, P. Tran (Ed.), Methods in Cell Biology, Elsevier, 2020, pp. 145–161. date_created: 2020-03-08T23:00:47Z date_published: 2020-02-27T00:00:00Z date_updated: 2026-04-08T07:26:30Z day: '27' department: - _id: MaLo doi: 10.1016/bs.mcb.2020.01.006 ec_funded: 1 editor: - first_name: 'Phong ' full_name: 'Tran, Phong ' last_name: Tran external_id: isi: - '000611826500008' intvolume: ' 158' isi: 1 language: - iso: eng main_file_link: - open_access: '1' url: https://doi.org/10.1101/839571 month: '02' oa: 1 oa_version: Preprint page: 145-161 project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell - _id: 260D98C8-B435-11E9-9278-68D0E5697425 name: Reconstitution of Bacterial Cell Division Using Purified Components publication: Methods in Cell Biology publication_identifier: issn: - 0091-679X publication_status: published publisher: Elsevier quality_controlled: '1' related_material: record: - id: '8358' relation: part_of_dissertation status: public scopus_import: '1' status: public title: Computational analysis of filament polymerization dynamics in cytoskeletal networks type: book_chapter user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87 volume: 158 year: '2020' ... --- _id: '7387' abstract: - lang: eng text: Most bacteria accomplish cell division with the help of a dynamic protein complex called the divisome, which spans the cell envelope in the plane of division. Assembly and activation of this machinery are coordinated by the tubulin-related GTPase FtsZ, which was found to form treadmilling filaments on supported bilayers in vitro1, as well as in live cells, in which filaments circle around the cell division site2,3. Treadmilling of FtsZ is thought to actively move proteins around the division septum, thereby distributing peptidoglycan synthesis and coordinating the inward growth of the septum to form the new poles of the daughter cells4. However, the molecular mechanisms underlying this function are largely unknown. Here, to study how FtsZ polymerization dynamics are coupled to downstream proteins, we reconstituted part of the bacterial cell division machinery using its purified components FtsZ, FtsA and truncated transmembrane proteins essential for cell division. We found that the membrane-bound cytosolic peptides of FtsN and FtsQ co-migrated with treadmilling FtsZ–FtsA filaments, but despite their directed collective behaviour, individual peptides showed random motion and transient confinement. Our work suggests that divisome proteins follow treadmilling FtsZ filaments by a diffusion-and-capture mechanism, which can give rise to a moving zone of signalling activity at the division site. acknowledgement: We acknowledge members of the Loose laboratory at IST Austria for helpful discussions—in particular, P. Caldas for help with the treadmilling analysis, M. Jimenez, A. Raso and N. Ropero for providing Alexa Fluor 488- and Alexa Fluor 647-labelled FtsA for the MST and analytical ultracentrifugation experiments. We thank C. You for providing the DODA-tris-NTA phospholipids, as well as J. Piehler and C. Richter (Department of Biology, University of Osnabruck, Germany) for the SLIMfast single-molecule tracking software and help with the confinement analysis. We thank J. Errington and H. Murray (both at Newcastle University, UK) for critical reading of the manuscript, and J. Brugués (MPI-CBG and MPI-PKS, Dresden, Germany) for help with the MATLAB programming and reading of the manuscript. This work was supported by the European Research Council through grant ERC-2015-StG-679239 to M.L. and grants HFSP LT 000824/2016-L4 and EMBO ALTF 1163-2015 to N.B., a grant from the Ministry of Economy and Competitiveness of the Spanish Government (BFU2016-75471-C2-1-P) to C.A. and G.R., and a Wellcome Trust Senior Investigator award (101824/Z/13/Z) and a grant from the BBSRC (BB/R017409/1) to W.V. article_processing_charge: No article_type: letter_note author: - first_name: Natalia S. full_name: Baranova, Natalia S. id: 38661662-F248-11E8-B48F-1D18A9856A87 last_name: Baranova orcid: 0000-0002-3086-9124 - first_name: Philipp full_name: Radler, Philipp id: 40136C2A-F248-11E8-B48F-1D18A9856A87 last_name: Radler orcid: '0000-0001-9198-2182 ' - first_name: Víctor M. full_name: Hernández-Rocamora, Víctor M. last_name: Hernández-Rocamora - first_name: Carlos full_name: Alfonso, Carlos last_name: Alfonso - first_name: Maria D full_name: Lopez Pelegrin, Maria D id: 319AA9CE-F248-11E8-B48F-1D18A9856A87 last_name: Lopez Pelegrin - first_name: Germán full_name: Rivas, Germán last_name: Rivas - first_name: Waldemar full_name: Vollmer, Waldemar last_name: Vollmer - first_name: Martin full_name: Loose, Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 citation: ama: Baranova NS, Radler P, Hernández-Rocamora VM, et al. Diffusion and capture permits dynamic coupling between treadmilling FtsZ filaments and cell division proteins. Nature Microbiology. 2020;5:407-417. doi:10.1038/s41564-019-0657-5 apa: Baranova, N. S., Radler, P., Hernández-Rocamora, V. M., Alfonso, C., Lopez Pelegrin, M. D., Rivas, G., … Loose, M. (2020). Diffusion and capture permits dynamic coupling between treadmilling FtsZ filaments and cell division proteins. Nature Microbiology. Springer Nature. https://doi.org/10.1038/s41564-019-0657-5 chicago: Baranova, Natalia S., Philipp Radler, Víctor M. Hernández-Rocamora, Carlos Alfonso, Maria D Lopez Pelegrin, Germán Rivas, Waldemar Vollmer, and Martin Loose. “Diffusion and Capture Permits Dynamic Coupling between Treadmilling FtsZ Filaments and Cell Division Proteins.” Nature Microbiology. Springer Nature, 2020. https://doi.org/10.1038/s41564-019-0657-5. ieee: N. S. Baranova et al., “Diffusion and capture permits dynamic coupling between treadmilling FtsZ filaments and cell division proteins,” Nature Microbiology, vol. 5. Springer Nature, pp. 407–417, 2020. ista: Baranova NS, Radler P, Hernández-Rocamora VM, Alfonso C, Lopez Pelegrin MD, Rivas G, Vollmer W, Loose M. 2020. Diffusion and capture permits dynamic coupling between treadmilling FtsZ filaments and cell division proteins. Nature Microbiology. 5, 407–417. mla: Baranova, Natalia S., et al. “Diffusion and Capture Permits Dynamic Coupling between Treadmilling FtsZ Filaments and Cell Division Proteins.” Nature Microbiology, vol. 5, Springer Nature, 2020, pp. 407–17, doi:10.1038/s41564-019-0657-5. short: N.S. Baranova, P. Radler, V.M. Hernández-Rocamora, C. Alfonso, M.D. Lopez Pelegrin, G. Rivas, W. Vollmer, M. Loose, Nature Microbiology 5 (2020) 407–417. corr_author: '1' date_created: 2020-01-28T16:14:41Z date_published: 2020-01-20T00:00:00Z date_updated: 2026-04-28T22:30:27Z day: '20' department: - _id: MaLo doi: 10.1038/s41564-019-0657-5 ec_funded: 1 external_id: isi: - '000508584700007' pmid: - '31959972' intvolume: ' 5' isi: 1 language: - iso: eng main_file_link: - open_access: '1' url: http://europepmc.org/article/PMC/7048620 month: '01' oa: 1 oa_version: Submitted Version page: 407-417 pmid: 1 project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell - _id: 259B655A-B435-11E9-9278-68D0E5697425 grant_number: LT000824/2016 name: Reconstitution of bacterial cell wall synthesis - _id: 2596EAB6-B435-11E9-9278-68D0E5697425 grant_number: ALTF 2015-1163 name: Synthesis of bacterial cell wall publication: Nature Microbiology publication_identifier: issn: - 2058-5276 publication_status: published publisher: Springer Nature quality_controlled: '1' related_material: link: - description: News on IST Homepage relation: press_release url: https://ist.ac.at/en/news/little-cell-big-cover-story/ record: - id: '14280' relation: dissertation_contains status: public scopus_import: '1' status: public title: Diffusion and capture permits dynamic coupling between treadmilling FtsZ filaments and cell division proteins type: journal_article user_id: 4359f0d1-fa6c-11eb-b949-802e58b17ae8 volume: 5 year: '2020' ... --- _id: '7197' abstract: - lang: eng text: During bacterial cell division, the tubulin-homolog FtsZ forms a ring-like structure at the center of the cell. This Z-ring not only organizes the division machinery, but treadmilling of FtsZ filaments was also found to play a key role in distributing proteins at the division site. What regulates the architecture, dynamics and stability of the Z-ring is currently unknown, but FtsZ-associated proteins are known to play an important role. Here, using an in vitro reconstitution approach, we studied how the well-conserved protein ZapA affects FtsZ treadmilling and filament organization into large-scale patterns. Using high-resolution fluorescence microscopy and quantitative image analysis, we found that ZapA cooperatively increases the spatial order of the filament network, but binds only transiently to FtsZ filaments and has no effect on filament length and treadmilling velocity. Together, our data provides a model for how FtsZ-associated proteins can increase the precision and stability of the bacterial cell division machinery in a switch-like manner. acknowledged_ssus: - _id: LifeSc - _id: Bio article_number: '5744' article_processing_charge: No article_type: original author: - first_name: Paulo R full_name: Dos Santos Caldas, Paulo R id: 38FCDB4C-F248-11E8-B48F-1D18A9856A87 last_name: Dos Santos Caldas orcid: 0000-0001-6730-4461 - first_name: Maria D full_name: Lopez Pelegrin, Maria D id: 319AA9CE-F248-11E8-B48F-1D18A9856A87 last_name: Lopez Pelegrin - first_name: Daniel J. G. full_name: Pearce, Daniel J. G. last_name: Pearce - first_name: Nazmi B full_name: Budanur, Nazmi B id: 3EA1010E-F248-11E8-B48F-1D18A9856A87 last_name: Budanur orcid: 0000-0003-0423-5010 - first_name: Jan full_name: Brugués, Jan last_name: Brugués - first_name: Martin full_name: Loose, Martin id: 462D4284-F248-11E8-B48F-1D18A9856A87 last_name: Loose orcid: 0000-0001-7309-9724 citation: ama: Dos Santos Caldas PR, Lopez Pelegrin MD, Pearce DJG, Budanur NB, Brugués J, Loose M. Cooperative ordering of treadmilling filaments in cytoskeletal networks of FtsZ and its crosslinker ZapA. Nature Communications. 2019;10. doi:10.1038/s41467-019-13702-4 apa: Dos Santos Caldas, P. R., Lopez Pelegrin, M. D., Pearce, D. J. G., Budanur, N. B., Brugués, J., & Loose, M. (2019). Cooperative ordering of treadmilling filaments in cytoskeletal networks of FtsZ and its crosslinker ZapA. Nature Communications. Springer Nature. https://doi.org/10.1038/s41467-019-13702-4 chicago: Dos Santos Caldas, Paulo R, Maria D Lopez Pelegrin, Daniel J. G. Pearce, Nazmi B Budanur, Jan Brugués, and Martin Loose. “Cooperative Ordering of Treadmilling Filaments in Cytoskeletal Networks of FtsZ and Its Crosslinker ZapA.” Nature Communications. Springer Nature, 2019. https://doi.org/10.1038/s41467-019-13702-4. ieee: P. R. Dos Santos Caldas, M. D. Lopez Pelegrin, D. J. G. Pearce, N. B. Budanur, J. Brugués, and M. Loose, “Cooperative ordering of treadmilling filaments in cytoskeletal networks of FtsZ and its crosslinker ZapA,” Nature Communications, vol. 10. Springer Nature, 2019. ista: Dos Santos Caldas PR, Lopez Pelegrin MD, Pearce DJG, Budanur NB, Brugués J, Loose M. 2019. Cooperative ordering of treadmilling filaments in cytoskeletal networks of FtsZ and its crosslinker ZapA. Nature Communications. 10, 5744. mla: Dos Santos Caldas, Paulo R., et al. “Cooperative Ordering of Treadmilling Filaments in Cytoskeletal Networks of FtsZ and Its Crosslinker ZapA.” Nature Communications, vol. 10, 5744, Springer Nature, 2019, doi:10.1038/s41467-019-13702-4. short: P.R. Dos Santos Caldas, M.D. Lopez Pelegrin, D.J.G. Pearce, N.B. Budanur, J. Brugués, M. Loose, Nature Communications 10 (2019). corr_author: '1' date_created: 2019-12-20T12:22:57Z date_published: 2019-12-17T00:00:00Z date_updated: 2026-04-08T07:26:30Z day: '17' ddc: - '570' department: - _id: MaLo - _id: BjHo doi: 10.1038/s41467-019-13702-4 ec_funded: 1 external_id: isi: - '000503009300001' file: - access_level: open_access checksum: a1b44b427ba341383197790d0e8789fa content_type: application/pdf creator: dernst date_created: 2019-12-23T07:34:56Z date_updated: 2020-07-14T12:47:53Z file_id: '7208' file_name: 2019_NatureComm_Caldas.pdf file_size: 8488733 relation: main_file file_date_updated: 2020-07-14T12:47:53Z has_accepted_license: '1' intvolume: ' 10' isi: 1 language: - iso: eng month: '12' oa: 1 oa_version: Published Version project: - _id: 2595697A-B435-11E9-9278-68D0E5697425 call_identifier: H2020 grant_number: '679239' name: Self-Organization of the Bacterial Cell - _id: 260D98C8-B435-11E9-9278-68D0E5697425 name: Reconstitution of Bacterial Cell Division Using Purified Components publication: Nature Communications publication_identifier: issn: - 2041-1723 publication_status: published publisher: Springer Nature quality_controlled: '1' related_material: record: - id: '8358' relation: dissertation_contains status: public scopus_import: '1' status: public title: Cooperative ordering of treadmilling filaments in cytoskeletal networks of FtsZ and its crosslinker ZapA tmp: image: /images/cc_by.png legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0) short: CC BY (4.0) type: journal_article user_id: c635000d-4b10-11ee-a964-aac5a93f6ac1 volume: 10 year: '2019' ...