Cytoskeletal and Actin‐Based Polymerization Motors and Their Role in Minimal Cell Design

Life implies motion. In cells, protein-based active molecular machines drive cell locomotion and intracellular transport, control cell shape, segregate genetic material, and split a cell in two parts. Key players among molecular machines driving these various cell functions are the cytoskeleton and motor proteins that convert chemical bound energy into mechanical work. Findings over the last decades in the field of in vitro reconstitutions of cytoskeletal and motor proteins have elucidated mechanistic details of these active protein systems. For example, a complex spatial and temporal interplay between the cytoskeleton and motor proteins is responsible for the translation of chemically bound energy into (directed) movement and force generation, which eventually governs the emergence of complex cellular functions. Understanding these mechanisms and the design principles of the cytoskeleton and motor proteins builds the basis for mimicking fundamental life processes. Here, a brief overview of actin, prokaryotic actin analogs, and motor proteins and their potential role in the design of a minimal cell from the bottom-up is provided

DNA Segregation in Natural and Synthetic Minimal Systems

Faithful segregation of replicated genomes to dividing daughter cells is a major hallmark of cellular life and needs to be part of the future design of the robustly proliferating minimal cell. So far, the complexity of eukaryotic chromosome segregation machineries has limited their applicability to synthetic systems. Prokaryotic plasmid segregation machineries offer promising alternative tools for bottom-up synthetic biology, with the first three-component DNA segregation system being reconstituted a decade ago. In this review, the mechanisms underlying DNA segregation in prokaryotes, with a particular focus on segregation of plasmids and chromosomal replication origins are reviewed, along with a brief discussion of archaeal and eukaryotic systems. In addition, this review shows how in vitro reconstitution has allowed deeper insights into these processes and discusses possible applications of these machineries for a minimal synthetic segrosome as well as the challenge of its coupling to a minimal replisome

Global monetary and financial spillovers: Evidence from a new measure of Bundesbank policy shocks

Identifying exogenous variation in monetary policy is crucial for investigating central bank policy transmission. Using newly-collected archival real-time data utilized by the Central Bank Council of the German Bundesbank, we identify unexpected changes in German monetary policy from 580 policy meetings between 1974 and 1998. German monetary policy shocks produce conventional effects on the German domestic economy: activity, prices, and credit decline significantly following a monetary contraction. But given Germany's central role in the European Monetary System (EMS), we can also shed light on debates about the international transmission of monetary policy and the relative importance of the U.S. Federal Reserve for the global cycle during these years. We find that Bundesbank policy spillovers were much stronger in major EMS economies with Deutschmark pegs than in non-EMS economies with floating exchange rates. Furthermore, compared to monetary spillovers from the U.S., German spillovers were comparable or even larger in magnitude for both pegs and floats

Functional Modules of Minimal Cell Division for Synthetic Biology

Cellular reproduction is one of the fundamental hallmarks of life. Therefore, the development of a minimal division machinery capable of proper genome condensation and organization, mid-cell positioning and segregation in space and time, and the final septation process constitute a fundamental challenge for synthetic biology. It is therefore important to be able to engineer such modules for the production of artificial minimal cells. A bottom-up assembly of molecular machines from bulk biochemicals complemented by in vivo experiments as well as computational modelling helps to approach such key cellular processes. Here, minimal functional modules involved in genome segregation and the division machinery and their spatial organization and positioning are reviewed, setting into perspective the design of a minimal cell. Furthermore, the milestones of recent in vitro reconstitution experiments in the context of cell division are discussed and their role in shedding light on fundamental cellular mechanisms that constitute spatiotemporal order is described. Lastly, current challenges in the field of bottom-up synthetic biology as well as possible future developments toward the development of minimal biomimetic systems are discussed

Reconstitution and coupling of DNA replication and segregation in a biomimetic system

A biomimetic system capable of replication and segregation of genetic material constitutes an essential component for the future design of a minimal synthetic cell. Here we have used the simple T7 bacteriophage system and the plasmid-derived ParMRC system to establish in vitro DNA replication and DNA segregation, respectively. These processes were incorporated into biomimetic compartments providing an enclosed reaction space. The functional lifetime of the encapsulated segregation system could be prolonged by equipping it with ATP-regenerating and oxygen-scavenging systems. Finally, we showed that DNA replication and segregation processes could be coupled in vitro by using condensed DNA nanoparticles resulting from DNA replication. ParM spindles extended over tens of micrometers and could thus be used for segregation in compartments that are significantly longer than bacterial cell size. Overall, this work demonstrates the successful bottom-up assembly and coupling of molecular machines that mediate replication and segregation, thus providing an important step towards the development of a fully functional minimal cell