TY - JOUR
T1 - Live Imaging of a Hyperthermophilic Archaeon Reveals Distinct Roles for Two ESCRT-III Homologs in Ensuring a Robust and Symmetric Division
AU - Pulschen, Andre Arashiro
AU - Mutavchiev, Delyan R.
AU - Culley, Siân
AU - Sebastian, Kim Nadine
AU - Roubinet, Jacques
AU - Roubinet, Marc
AU - Risa, Gabriel Tarrason
AU - van Wolferen, Marleen
AU - Roubinet, Chantal
AU - Schmidt, Uwe
AU - Dey, Gautam
AU - Albers, Sonja Verena
AU - Henriques, Ricardo
AU - Baum, Buzz
N1 - Funding Information:
The authors would like to thank the entire Baum lab for their input throughout the project. We would like to thank Juan Manuel Garcia-Arcos for help early on in this project; Tobias Härtel, Pedro Matos Pereira, and Alexandre Bisson for discussions about fluorescent probes; Alexander Wagner for providing the plasmid pSVAaraFX-stop; Alexandre Bisson and Jan Löwe for comments on the manuscript; and Mateusz Trylinski for helping with movies. A.A.P. was supported by the HFSP LT001027/2019 fellowship. D.R.M. was supported by the BBSRC ( BB/K009001/1 ). G.T.R. was supported by the MRC PhD studentship award ( MC_CF12266 ). G.D. was funded by a European Union Marie Sklodowska-Curie Individual Fellowship ( 704281-CCDSA ). U.S. was supported by the BMBF grant SYSBIO II ( 031L0044 ). In addition, A.A.P., S.C., and G.D. all received generous support from the Wellcome Trust ( 203276/Z/16/Z ). Research was supported by Wellcome Trust ( 203276/Z/16/Z ) and Volkswagen Foundation ( Az 96727 ) with additional support for B.B. and R.H. provided by UCL and the MRC ( MC_CF12266 ).
Publisher Copyright:
© 2020 The Authors
PY - 2020/7/20
Y1 - 2020/7/20
N2 - Live-cell imaging has revolutionized our understanding of dynamic cellular processes in bacteria and eukaryotes. Although similar techniques have been applied to the study of halophilic archaea [1–5], our ability to explore the cell biology of thermophilic archaea has been limited by the technical challenges of imaging at high temperatures. Sulfolobus are the most intensively studied members of TACK archaea and have well-established molecular genetics [6–9]. Additionally, studies using Sulfolobus were among the first to reveal striking similarities between the cell biology of eukaryotes and archaea [10–15]. However, to date, it has not been possible to image Sulfolobus cells as they grow and divide. Here, we report the construction of the Sulfoscope, a heated chamber on an inverted fluorescent microscope that enables live-cell imaging of thermophiles. By using thermostable fluorescent probes together with this system, we were able to image Sulfolobus acidocaldarius cells live to reveal tight coupling between changes in DNA condensation, segregation, and cell division. Furthermore, by imaging deletion mutants, we observed functional differences between the two ESCRT-III proteins implicated in cytokinesis, CdvB1 and CdvB2. The deletion of cdvB1 compromised cell division, causing occasional division failures, whereas the ΔcdvB2 exhibited a profound loss of division symmetry, generating daughter cells that vary widely in size and eventually generating ghost cells. These data indicate that DNA separation and cytokinesis are coordinated in Sulfolobus, as is the case in eukaryotes, and that two contractile ESCRT-III polymers perform distinct roles to ensure that Sulfolobus cells undergo a robust and symmetrical division.
AB - Live-cell imaging has revolutionized our understanding of dynamic cellular processes in bacteria and eukaryotes. Although similar techniques have been applied to the study of halophilic archaea [1–5], our ability to explore the cell biology of thermophilic archaea has been limited by the technical challenges of imaging at high temperatures. Sulfolobus are the most intensively studied members of TACK archaea and have well-established molecular genetics [6–9]. Additionally, studies using Sulfolobus were among the first to reveal striking similarities between the cell biology of eukaryotes and archaea [10–15]. However, to date, it has not been possible to image Sulfolobus cells as they grow and divide. Here, we report the construction of the Sulfoscope, a heated chamber on an inverted fluorescent microscope that enables live-cell imaging of thermophiles. By using thermostable fluorescent probes together with this system, we were able to image Sulfolobus acidocaldarius cells live to reveal tight coupling between changes in DNA condensation, segregation, and cell division. Furthermore, by imaging deletion mutants, we observed functional differences between the two ESCRT-III proteins implicated in cytokinesis, CdvB1 and CdvB2. The deletion of cdvB1 compromised cell division, causing occasional division failures, whereas the ΔcdvB2 exhibited a profound loss of division symmetry, generating daughter cells that vary widely in size and eventually generating ghost cells. These data indicate that DNA separation and cytokinesis are coordinated in Sulfolobus, as is the case in eukaryotes, and that two contractile ESCRT-III polymers perform distinct roles to ensure that Sulfolobus cells undergo a robust and symmetrical division.
KW - archaea
KW - cell division
KW - DNA segregation
KW - ESCRT-III
KW - hyperthermophiles
KW - live-cell imaging
KW - Sulfolobus acidocaldarius
KW - thermophiles
UR - http://www.scopus.com/inward/record.url?scp=85086646974&partnerID=8YFLogxK
U2 - 10.1016/j.cub.2020.05.021
DO - 10.1016/j.cub.2020.05.021
M3 - Article
C2 - 32502411
AN - SCOPUS:85086646974
SN - 0960-9822
VL - 30
SP - 2852-2859.e4
JO - Current Biology
JF - Current Biology
IS - 14
ER -