Archaeal roots of eukaryotic cell cycle control and ESCRT-III mediated cell division

The life we see around us is predominantly eukaryotic, made of large multicellular organisms such as fungi, plants, and animals. However, all eukaryotes likely evolved more than a billion years ago from a gradual symbiotic merger of an archaeon and a bacterium. Since this time, eukaryotes have evolved and diversified, but certain fundamental features of their biology can be traced back to their archaeal-bacterial origins. In my PhD I have sought to apply this evolutionary perspective to shed light on universal principles of cellular growth and division by studying these processes in \textit{Sulfolobus acidocaldarius}, one of the closest relatives of eukaryotes that is also experimentally tractable. I focussed on the cell cycle, which in eukaryotes is marked by tightly controlled discrete cycles of DNA replication and cell division. These features are shared by \textit{S. acidocaldarius}. Progress through the eukaryotic cell cycle is controlled by cyclin-dependent kinases, whose activities oscillate due to coordinated waves of expression and proteasome-mediated degradation of cyclins. Contrastingly, neither \textit{S. acidocadarius} nor any other archaea known to date possess cyclins or cyclin-dependent kinases. In searching for other regulatory principles shared by the archaeal and eukaryotic cell cycle, we observed that \textit{S. acidocaldarius} cells use a pattern of oscillating gene expression and targeted proteasome-mediated degradation to complete their cell cycle – just as eukaryotes do. Specifically, we were able to show that cell constriction is triggered by targeted proteasome-mediated degradation of CdvB, an ESCRT-III homologue expressed in the run-up to cell division. These findings further support the idea that the eukaryotic cell cycle control has its origins in archaea and provide insight into how a minimal ESCRT-III system can cut the membrane

The patterned assembly and stepwise Vps4-mediated disassembly of composite ESCRT-III polymers drives archaeal cell division

ESCRT-III family proteins form composite polymers that deform and cut membrane tubes in the context of a wide range of cell biological processes across the tree of life. In reconstituted systems, sequential changes in the composition of ESCRT-III polymers induced by the AAA-adenosine triphosphatase Vps4 have been shown to remodel membranes. However, it is not known how composite ESCRT-III polymers are organized and remodeled in space and time in a cellular context. Taking advantage of the relative simplicity of the ESCRT-III-dependent division system in Sulfolobus acidocaldarius, one of the closest experimentally tractable prokaryotic relatives of eukaryotes, we use super-resolution microscopy, electron microscopy, and computational modeling to show how CdvB/CdvB1/CdvB2 proteins form a precisely patterned composite ESCRT-III division ring, which undergoes stepwise Vps4-dependent disassembly and contracts to cut cells into two. These observations lead us to suggest sequential changes in a patterned composite polymer as a general mechanism of ESCRT-III-dependent membrane remodeling. </p

The proteasome controls ESCRT-III–mediated cell division in an archaeon

INTRODUCTION: Eukaryotes likely arose from a symbiotic partnership between an archaeal host and an alpha-proteobacterium, giving rise to the cell body and the mitochondria, respectively. Because of this, a number of proteins controlling key events in the eukaryotic cell division cycle have their origins in archaea. These include ESCRT-III proteins, which catalyze the final step of cytokinesis in many eukaryotes and in the archaeon Sulfolobus acidocaldarius. However, to date, no archaeon has been found that harbors homologs of cell cycle regulators, like cyclin-dependent kinases and cyclins, which order events in the cell cycle across all eukaryotes. Thus, it remains uncertain how key events in the archaeal cell cycle, including division, are regulated. RATIONALE: An exception to this is the 20S proteasome, which is conserved between archaea and eukaryotes and which regulates the eukaryotic cell cycle through the degradation of cyclins. To explore the function of the 20S proteasome in the archaeon S. acidocaldarius, we determined its structure by crystallography and carried out in vitro biochemical analyses of its activity with and without inhibition. The impact of proteasome inhibition on cell division and cell cycle progression was examined in vivo by flow cytometry and super-resolution microscopy. Following up with mass spectrometry, we identified proteins degraded by the proteasome during division. Finally, we used molecular dynamics simulations to model the mechanics of this process. RESULTS: Here, we present a structure of the 20S proteasome of S. acidocaldarius to a resolution of 3.7 Å, which we used to model its sensitivity to the eukaryotic inhibitor bortezomib. When this inhibitor was added to synchronous cultures, it was found to arrest cells mid-division, with a stable ESCRT-III division ring positioned at the cell center between the two separated and prereplicative nucleoids. Proteomics was then used to identify a single archaeal ESCRT-III homolog, CdvB, as a key target of the proteasome that must be degraded to enable division to proceed. Examining the localization patterns of CdvB and two other archaeal ESCRT-III homologs, CdvB1 and CdvB2, by flow cytometry and super-resolution microscopy revealed the sequence of events that leads to division. First, a CdvB ring is assembled. This CdvB ring then templates the assembly of the contractile ESCRT-III homologs, CdvB1 and CdvB2, to form a composite division ring. Cell division is then triggered by proteasome-mediated degradation of CdvB, which allows the CdvB1:CdvB2 copolymer to constrict, pulling the membrane with it. During constriction, the CdvB1:CdvB2 copolymer is disassembled, thus vacating the membrane neck to drive abscission, yielding two daughter cells with diffuse CdvB1 and CdvB2. CONCLUSION: This study reveals a role for the proteasome in driving structural changes in a composite ESCRT-III copolymer, enabling the stepwise assembly, disassembly, and contraction of an ESCRT-III–based division ring. Although it is not yet clear how proteasomal inhibition prevents S. acidocaldarius cells from resetting the cell cycle to initiate the next S phase, these data strengthen the case for the eukaryotic cell cycle regulation having its origins in archaea