Abstract
Cell division is fundamental to all cellular life. Most archaea depend on either the prokaryotic tubulin homologue FtsZ or the endosomal sorting complex required for transport for division but neither system has been robustly characterized. Here, we show that three of the four photosynthesis reaction centre barrel domain proteins of Haloferax volcanii (renamed cell division proteins B1/2/3 (CdpB1/2/3)) play important roles in cell division. CdpB1 interacts directly with the FtsZ membrane anchor SepF and is essential for cell division, whereas deletion of cdpB2 and cdpB3 causes a major and a minor division defect, respectively. Orthologues of CdpB proteins are also involved in cell division in other haloarchaea, indicating a conserved function of these proteins. Phylogenetic analysis shows that photosynthetic reaction centre barrel proteins are widely distributed among archaea and appear to be central to cell division in most if not all archaea.
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Data availability
Data generated and analysed during this study are presented in the text or in the Supplementary Information and Supplementary Data. Plasmids and strains that support the findings of this study are available from the corresponding authors. Source data are provided with this paper.
References
Wang, X. & Lutkenhaus, J. FtsZ ring: the eubacterial division apparatus conserved in archaebacteria. Mol. Microbiol. 21, 313–319 (1996).
Samson, R. Y., Obita, T., Freund, S. M., Williams, R. L. & Bell, S. D. A role for the ESCRT system in cell division in archaea. Science 322, 1710–1713 (2008).
Lindas, A. C., Karlsson, E. A., Lindgren, M. T., Ettema, T. J. & Bernander, R. A unique cell division machinery in the archaea. Proc. Natl Acad. Sci. USA 105, 18942–18946 (2008).
Makarova, K. S., Yutin, N., Bell, S. D. & Koonin, E. V. Evolution of diverse cell division and vesicle formation systems in archaea. Nat. Rev. Microbiol. 8, 731–741 (2010).
Caspi, Y. & Dekker, C. Dividing the archaeal way: the ancient Cdv cell-division machinery. Front. Microbiol. 9, 174 (2018).
Ithurbide, S., Gribaldo, S., Albers, S. V. & Pende, N. Spotlight on FtsZ-based cell division in archaea. Trends Microbiol. 30, 665–678 (2022).
van Wolferen, M., Pulschen, A. A., Baum, B., Gribaldo, S. & Albers, S. V. The cell biology of archaea. Nat. Microbiol. 7, 1744–1755 (2022).
Margolin, W., Wang, R. & Kumar, M. Isolation of an ftsZ homolog from the archaebacterium Halobacterium salinarium: implications for the evolution of FtsZ and tubulin. J. Bacteriol. 178, 1320–1327 (1996).
Baumann, P. & Jackson, S. P. An archaebacterial homologue of the essential eubacterial cell division protein FtsZ. Proc. Natl Acad. Sci. USA 93, 6726–6730 (1996).
Blanch Jover, A. & Dekker, C. The archaeal Cdv cell division system. Trends Microbiol. 31, 601–615 (2023).
Liao, Y., Ithurbide, S., Evenhuis, C., Lowe, J. & Duggin, I. G. Cell division in the archaeon Haloferax volcanii relies on two FtsZ proteins with distinct functions in division ring assembly and constriction. Nat. Microbiol. 6, 594–605 (2021).
Nussbaum, P., Gerstner, M., Dingethal, M., Erb, C. & Albers, S. V. The archaeal protein SepF is essential for cell division in Haloferax volcanii. Nat. Commun. 12, 3469 (2021).
Pende, N. et al. SepF is the FtsZ anchor in archaea, with features of an ancestral cell division system. Nat. Commun. 12, 3214 (2021).
Anantharaman, V. & Aravind, L. The PRC-barrel: a widespread, conserved domain shared by photosynthetic reaction center subunits and proteins of RNA metabolism. Genome Biol. 3, RESEARCH0061 (2002).
Liao, Y. et al. CdrS is a global transcriptional regulator influencing cell division in Haloferax volcanii. mBio 12, e0141621 (2021).
Darnell, C. L. et al. The ribbon-helix-helix domain protein CdrS regulates the tubulin homolog ftsZ2 to control cell division in Archaea. mBio https://doi.org/10.1128/mBio.01007-20 (2020).
Allers, T., Ngo, H. P., Mevarech, M. & Lloyd, R. G. Development of additional selectable markers for the halophilic archaeon Haloferax volcanii based on the leuB and trpA genes. Appl. Environ. Microbiol. 70, 943–953 (2004).
Large, A. et al. Characterization of a tightly controlled promoter of the halophilic archaeon Haloferax volcanii and its use in the analysis of the essential cct1 gene. Mol. Microbiol. 66, 1092–1106 (2007).
Hu, C. D. & Kerppola, T. K. Simultaneous visualization of multiple protein interactions in living cells using multicolor fluorescence complementation analysis. Nat. Biotechnol. 21, 539–545 (2003).
Cabantous, S. et al. A new protein-protein interaction sensor based on tripartite split-GFP association. Sci. Rep. 3, 2854 (2013).
Tehrani, A., Prince, R. C. & Beatty, J. T. Effects of photosynthetic reaction center H protein domain mutations on photosynthetic properties and reaction center assembly in Rhodobacter sphaeroides. Biochemistry 42, 8919–8928 (2003).
Lovgren, J. M. et al. The PRC-barrel domain of the ribosome maturation protein RimM mediates binding to ribosomal protein S19 in the 30S ribosomal subunits. RNA 10, 1798–1812 (2004).
Abecasis, A. B. et al. A genomic signature and the identification of new sporulation genes. J. Bacteriol. 195, 2101–2115 (2013).
Samson, R. Y. et al. Molecular and structural basis of ESCRT-III recruitment to membranes during archaeal cell division. Mol. Cell 41, 186–196 (2011).
Moriscot, C. et al. Crenarchaeal CdvA forms double-helical filaments containing DNA and interacts with ESCRT-III-like CdvB. PLoS ONE 6, e21921 (2011).
Weiss, C. A. et al. NrnA is a 5′-3′ exonuclease that processes short RNA substrates in vivo and in vitro. Nucleic Acids Res. 50, 12369–12388 (2022).
Chen, A. W. et al. The role of 3′ to 5′ reverse RNA polymerization in tRNA fidelity and repair. Genes 10, 250 (2019).
Moir, D., Stewart, S. E., Osmond, B. C. & Botstein, D. Cold-sensitive cell-division-cycle mutants of yeast: isolation, properties and pseudoreversion studies. Genetics 100, 547–563 (1982).
Dassain, M., Leroy, A., Colosetti, L., Carole, S. & Bouche, J. P. A new essential gene of the ‘minimal genome’ affecting cell division. Biochimie 81, 889–895 (1999).
Nußbaum, P. et al. PRC domain-containing proteins modulate FtsZ-based archaeal cell division. Preprint at bioRxiv https://doi.org/10.1101/2023.03.28.534543 (2023).
Maupin-Furlow, J. Proteasomes and protein conjugation across domains of life. Nat. Rev. Microbiol. 10, 100–111 (2011).
Duggin, I. G. et al. CetZ tubulin-like proteins control archaeal cell shape. Nature 519, 362–365 (2015).
de Silva, R. T. et al. Improved growth and morphological plasticity of Haloferax volcanii. Microbiology https://doi.org/10.1099/mic.0.001012 (2021).
Ye, X., Ou, J., Ni, L., Shi, W. & Shen, P. Characterization of a novel plasmid from extremely halophilic archaea: nucleotide sequence and function analysis. FEMS Microbiol. Lett. 221, 53–57 (2003).
Liu, H., Han, J., Liu, X., Zhou, J. & Xiang, H. Development of pyrF-based gene knockout systems for genome-wide manipulation of the archaea Haloferax mediterranei and Haloarcula hispanica. J. Genet. Genomics 38, 261–269 (2011).
Wang, J. et al. A novel family of tyrosine integrases encoded by the temperate pleolipovirus SNJ2. Nucleic Acids Res. 46, 2521–2536 (2018).
Allers, T., Barak, S., Liddell, S., Wardell, K. & Mevarech, M. Improved strains and plasmid vectors for conditional overexpression of His-tagged proteins in Haloferax volcanii. Appl. Environ. Microbiol. 76, 1759–1769 (2010).
Cai, S. et al. Identification of the haloarchaeal phasin (PhaP) that functions in polyhydroxyalkanoate accumulation and granule formation in Haloferax mediterranei. Appl. Environ. Microbiol. 78, 1946–1952 (2012).
de Boer, P. A., Crossley, R. E. & Rothfield, L. I. A division inhibitor and a topological specificity factor coded for by the minicell locus determine proper placement of the division septum in E. coli. Cell 56, 641–649 (1989).
Bernhardt, T. G. & de Boer, P. A. SlmA, a nucleoid-associated, FtsZ binding protein required for blocking septal ring assembly over chromosomes in E. coli. Mol. Cell 18, 555–564 (2005).
Hale, C. A. & de Boer, P. A. Direct binding of FtsZ to ZipA, an essential component of the septal ring structure that mediates cell division in E. coli. Cell 88, 175–185 (1997).
Pichoff, S. & Lutkenhaus, J. Identification of a region of FtsA required for interaction with FtsZ. Mol. Microbiol. 64, 1129–1138 (2007).
Du, S. & Lutkenhaus, J. SlmA antagonism of FtsZ assembly employs a two-pronged mechanism like MinCD. PLoS Genet. 10, e1004460 (2014).
Makarova, K. S., Wolf, Y. I. & Koonin, E. V. Archaeal clusters of orthologous genes (arCOGs): an update and application for analysis of shared features between Thermococcales, Methanococcales and Methanobacteriales. Life 5, 818–840 (2015).
Makarova, K. S., Wolf, Y. I. & Koonin, E. V. Towards functional characterization of archaeal genomic dark matter. Biochem. Soc. Trans. 47, 389–398 (2019).
Altschul, S. F. et al. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25, 3389–3402 (1997).
Zimmermann, L. et al. A completely reimplemented MPI Bioinformatics Toolkit with a new HHpred server at its core. J. Mol. Biol. 430, 2237–2243 (2018).
Edgar, R. C. Muscle5: high-accuracy alignment ensembles enable unbiased assessments of sequence homology and phylogeny. Nat. Commun. 13, 6968 (2022).
Esterman, E. S., Wolf, Y. I., Kogay, R., Koonin, E. V. & Zhaxybayeva, O. Evolution of DNA packaging in gene transfer agents. Virus Evol. 7, veab015 (2021).
Price, M. N., Dehal, P. S. & Arkin, A. P. FastTree 2–approximately maximum-likelihood trees for large alignments. PLoS ONE 5, e9490 (2010).
Delorenzi, M. & Speed, T. An HMM model for coiled-coil domains and a comparison with PSSM-based predictions. Bioinformatics 18, 617–625 (2002).
Acknowledgements
We thank members of the Du laboratory, Koonin laboratory, Chen laboratory and Krupovic laboratory for advice and helpful discussions to carry out this study. We thank Y. Liao and I. Duggin at University of Technology, Sydney, for sending us the ftsZ depletion/deletion strains and plasmids for construction of fluorescent protein fusions. We thank T. Allers at University of Nottingham and X. Liu at Shanghai Jiao Tong University for sending us the H. volcanii strains and plasmids. We thank M. Li and H. Xiang at the Institute of Microbiology Chinese Academy of Sciences for providing us with the H. hispanica strains and plasmids. We would also like to thank J. Liu and Y. Yang at Shandong University for insightful discussions on the function of the PRC barrel domain of CdvA. This study was supported by National Natural Science Foundation of China (grant nos. 32270049 and 32070032, http://www.nsfc.gov.cn/), the Fundamental Research Funds for the Central Universities (grant no. 2042021kf0198) and Wuhan University (https://www.whu.edu.cn/) to S.D.; the research of K.S.M. and E.V.K. is supported by the Intramural Research Program at the National Library of Medicine, National Institute of Health, USA.
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S.D., S.Z. and E.V.K. conceptualized this study. S.Z., K.S.M., W.Z., Y.L., L.Z., Q.W. and H.G. developed the methodology and carried out the investigations and acquired the data. S.D., S.Z., K.S.M., X.C., J.L. M.K. and E.V.K. interpreted the data. S.D., X.C., K.S.M. and E.V.K. obtained resources. S.D., X.C., J.L. and E.V.K. undertook supervision. S.D., S.Z., K.S.M., M.K., X.C., J.L. and E.V.K. wrote the original draft and reviewed and edited the final manuscript.
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Extended data
Extended Data Fig. 1 CdpB1 does not depend on FtsZ1 for colocalization with FtsZ2 and SepF.
a. Representative images of CdpB1-GFP and FtsZ2-mCherry localization in the presence or absence of FtsZ1, respectively. b. Representative images of CdpB1-GFP and SepF-mCherry localization in the presence or absence of FtsZ1, respectively. Depletion of FtsZ1 was achieved by removal of tryptophan from the cultures. Strain ID56 (H98, Ptna::ftsZ1) harbouring plasmid pZS284 (PphaR::cdpB1-gfp-ftsZ2-mCherry) or pZS285 (PphaR::cdpB1-gfp-sepF-mCherry) were grown in Hv-Cab medium with or without tryptophan. Percentage of protein colocalization (%) were indicated, number of GFP/mCherry structures or foci (n > 200). Scale bars 5 μm.
Extended Data Fig. 2 CdpB1 does not depend on FtsZ2 for colocalization with FtsZ1 and SepF.
a. Representative images of CdpB1-GFP and FtsZ1-mCherry localization in the presence or absence of FtsZ2, respectively. b. Representative images of CdpB1-GFP and SepF-mCherry localization in the presence or absence of FtsZ2, respectively. Depletion of FtsZ2 was achieved by removal of tryptophan from the cultures. Strain ID57 (H98, Ptna::ftsZ2) harbouring plasmid pZS239 (PphaR::cdpB1-gfp-ftsZ1-mCherry) or pZS285 (PphaR::cdpB1-gfp-sepF-mCherry) were grown in Hv-Cab medium with or without tryptophan. White arrows indicate the colocalization of the GFP and mCherry fluorescence signal, cyan arrows indicate the CdpB1-GFP foci or aggregates. Percentage of protein colocalization (%) were indicated, number of GFP/mCherry structures or foci (n > 200). Scale bars 5 μm.
Extended Data Fig. 3 CdpB1 depends on SepF for correct localization.
a. Representative images of CdpB1-GFP and FtsZ2-mCherry localization in the presence or absence of SepF, respectively. Depletion of SepF was achieved by removal of tryptophan from the cultures. Strain HZS2 (H98, Ptna::sepF) harbouring plasmid pZS284 (PphaR:: cdpB1-gfp-ftsZ2-mCherry) were grown in Hv-Cab medium with or without tryptophan. White arrows indicate the colocalization of the GFP and mCherry fluorescence signal, cyan arrows indicate the CdpB1-GFP foci or aggregates. Percentage of protein colocalization (%) were indicated, number of GFP/mCherry structures or foci (n > 200). Scale bars 5 μm.
Extended Data Fig. 4 Determination of the level of CdpB1 necessary for complementation.
a-b. CdpB1-depleted cells resume normal cell shape and size following restoration of CdpB1 expression. HZS1 (H98, Ptna::cdpB1) or HZS24 (H98, Ptna::his-cdpB1) grown in Hv-Cab (+50 μg/mL uracil) without tryptophan for 24 hours was diluted 1:100 in fresh medium with indicated concentrations of tryptophan. 15 hours later, samples were spotted onto a BSW agarose pad for microscopy. Scale bars 5 μm. c. Western blot to check the level of His-CdpB1 at the indicated concentration of tryptophan. Samples from panel b were prepared for SDS–PAGE and Western blot. The level of His-CdpB1 increased as the concentration of tryptophan in the medium increased. Strain H98, which did not express any His-tagged protein, was run as a negative control. Upper panel, Western blot; lower panel, Coomassie brilliant blue staining.
Extended Data Fig. 5 Cell morphology and protein localization upon depletion and repletion of division proteins.
a-c. Representative images of FtsZ1-GFP localization and cell morphology after depletion of FtsZ2 (a), SepF (b) or CdpB1 (c). Strains ID57 (H98,Ptna::ftsZ2), HZS2 (H98, Ptna::sepF) or HZS1 (H98, Ptna::cdpB1) harbouring plasmid pZS208 (Pnative:: ftsZ1-gfp) were grown in Hv-Cab medium without tryptophan to deplete CdpB1. Samples were taken at the indicated time points after the removal of tryptophan. Red arrows indicate multiple spiral and large abnormal FtsZ1-GFP localization in rod-shaped and large cells. Scale bars 5 μm. d-f. Representative images of FtsZ1-GFP (d), FtsZ2-GFP (e) and SepF-GFP (f) localization and cell morphology in CdpB1 repleted cells. Strain HZS1 (H98, Ptna::cdpB1) harbouring plasmids expressing fluorescent division protein fusions was grown in Hv-Cab medium without tryptophan for 24 hours and then inoculated into fresh Hv-Cab medium with 1 mM tryptophan. Samples were taken at the indicated time points after the addition of tryptophan. Red arrows indicate the nearly normal localization of FtsZ1-GFP, FtsZ2-GFP and SepF-GFP 9 hours after the addition of tryptophan to induce CdpB1. Scale bars 5 μm.
Extended Data Fig. 6 CdpB1 displays interaction with FtsZ1 and FtsZ2 in Split-FP assay.
a. Representative images of Split-FP assay showing the interaction signal between CdpB1 and SepF. b. Representative images of Split-FP assay showing the interaction signal between CdpB1 and FtsZ2. c. Representative images of Split-FP assay showing the interaction signal between CdpB1 and FtsZ1. An exponential culture of strain H26 (DS70, ΔpyrE2) harbouring the Split-FP plasmid expressing the indicated protein(s) was treated as in Fig. 3a and the fluorescence signal and localization were observed by microscopy. Scale bars 5 μm.
Extended Data Fig. 7 Co-IP experiments show that CdpB1 does not interact with FtsZ1 and FtsZ2.
The experiment was carried out as in Fig. 3c. a. CdpB1-His did not co-immunoprecipitated with FtsZ1-GFP. b. CdpB1-His did not co-immunoprecipitated with FtsZ2-GFP.
Extended Data Fig. 8 Alignment of PRC barrel proteins and their predicted structures.
a. Amino acid sequences of PRC barrel domain containing proteins from H. volcanii, Natrinema sp. J7-1 and H. hispanica were downloaded from Uniprot: https://www.uniprot.org/, aligned by Clustal Omega: https://www.ebi.ac.uk/Tools/msa/clustalo/ and then depicted using ESPRIPT 3.0: http://espript.ibcp.fr/. b. Phylogenetic tree of the PRC barrel domain containing proteins from H. volcanii, Natrinema sp. J7-1 and H. hispanica generated by Clustal Omega. c. Superimposition of the predicted structures of CdpB1, CdpB2, CdpB3 and HVO_1607 of H. volcanii. Structure models were generated by AlphaFold and were downloaded from Uniprot and aligned by PyMOL. CdpB1, green; CdpB2, cyan; CdpB3, magenta; HVO_1607, yellow.
Extended Data Fig. 9 Interaction between the CdpB proteins in vivo.
a-c. Interaction between the CdpB proteins determined by Split-FP assay. a. Representative images showing the interaction signal between CdpB1 and CdpB2. b. Representative images showing the interaction signal between CdpB2 and CdpB3. c. Representative images showing the interaction signal between CdpB1 and CdpB3. An exponential phase culture of strain H26 (DS70, ΔpyrE2) harbouring the Split-FP plasmid expressing the indicated protein(s) was treated as in Fig. 3a and the fluorescence signal and localization were examined by microscopy. Scale bars 5 μm. d-f. Co-IP experiments to determine the interactions between CdpB proteins. d. CdpB1-His and CdpB2-GFP immunoprecipitated with each other. e. CdpB2-His and CdpB3-GFP immunoprecipitated with each other. f. CdpB1-His and CdpB3-GFP did not immunoprecipitate with each other. The experiment was carried out as in Fig. 3c. Supernatants were incubated with rabbit antibodies coated magnetic beads, while the following Western blotting analysis used mouse antibodies.
Extended Data Fig. 10 Localization dependence of PRC barrel proteins in Natrinema sp. J7 and comparison of the PRC barrel domains of CdpB1 and CdvA from S. acidocaldricus.
a. Midcell localization of NJ7G_2779 and NJ7G_3497 depends on NJ7GCdpB1 in Natrinema sp. J7. Strain HZS4 (CJ7-F, Ptna::NJ7GcdpB1) carrying plasmid pZS513 (Pnative::NJ7G_2729-gfp) or pZS514 (Pnative::NJ7G_3497-gfp) was grown in Hv-Cab medium with or without tryptophan to check cell morphology and protein localization. White arrows indicate the midcell localization of NJ7G_3479-GFP. Scale bars 5 μm. b. AlphaFold structural models of H. volcanii CdpB1 and CdvA from S. acidocaldricus. Structural models of CdpB1 (D4H036) and CdvA (Q4J923) were downloaded from Uniprot and aligned with PyMOL.
Supplementary information
Supplementary Information
Supplementary Figs. 1–4 and Note.
Supplementary Data 1 and 3–7
Supplementary Data 1, Complete list of PRC barrel domain proteins for genomes from arCOG database. Data 3, PRC barrel genes neighbourhoods. Data 4, Strains used in this study. Data 5, Plasmids used in this study. Data 6, Primers used in this study. Data 7, Reagents and chemicals used in this study.
Supplementary Data 2
The complete phylogenetic tree of PRC barrel genes in Newick format.
Source Data for Supplementary Fig. 3c
Raw data for RT–qPCR.
Source Data for Supplementary Fig. 3d
OD600 of WT and CdpB1 depletion strains growing in the presence and absence of tryptophan.
Source data
Source Data Fig. 1e
Unprocessed western blot and gel for Fig. 1e.
Source Data Fig. 3c,d
Unprocessed western blots for Fig. 3c,d.
Source Data Extended Data Fig. 4c
Unprocessed western blots for Extended Data Fig. 4c.
Source Data Extended Data Fig. 7a,b
Unprocessed western blot and gel for Extended Data Fig. 7a,b.
Source Data Extended Data Fig. 9d–f
Unprocessed western blots for Extended Data Fig. 9d–f.
Source Data Fig. 3b
Statistical source data for Fig. 3b.
Source Data Colocalization
Statistical source data for protein colocalization.
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Zhao, S., Makarova, K.S., Zheng, W. et al. Widespread photosynthesis reaction centre barrel proteins are necessary for haloarchaeal cell division. Nat Microbiol 9, 712–726 (2024). https://doi.org/10.1038/s41564-024-01615-y
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DOI: https://doi.org/10.1038/s41564-024-01615-y
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