Biology subjects

Greiss, F.

Publications and source records attributed to Greiss, F..

4 recordsLinked to original sources

bioRxiv2024

Transcription swells chromosomes in vitro

Transplanting chromosomes from living to artificial cells would impact our understanding of chromosome organization and DNA transactions, with implications for autonomous biological systems. Here, we transplanted Escherichia coli chromosomes into artificial cells, enabling real-time labeling, manipulation, and steady-state gene expression down to the single-molecule limit. Chromosomes stripped of native proteins transitioned from a swollen to compacted state induced by transcription inhibition, in contrast to protein-bound chromosomes retaining an organization with blobs. In a cell-free expression reaction, RNA polymerases were uniformly distributed along the entire chromosome and rapidly detached, consistent with a global transcriptional activity. We used tailored surfaces to capture and count 20 nascent proteins per hour from a single gene on the chromosome. We mapped stably bound condensins to the blobs, supporting a model where swelling by transcription is counterbalanced by condensin-mediated compaction. Our data suggest transplanted chromosomes as active gels organized by molecular machines.

biophysics↗

bioRxiv2023

A genetic circuit on a single DNA molecule as autonomous dissipative nanodevice

Realizing genetic circuits on single DNA molecules as self-encoded dissipative nanodevices is a major step toward miniaturization of autonomous biological systems. A circuit operating on a single DNA implies that genetically encoded proteins localize during coupled transcriptiontranslation to DNA, but a single-molecule measurement demonstrating this has remained a challenge. Here, we used a genetically encoded fluorescent reporter system with improved spatiotemporal resolution and observed the synthesis of individual proteins tethered to a DNA molecule by transient complexes of RNA polymerase, messenger RNA, and ribosome. Against expectations in dilute cell-free conditions where equilibrium considerations favor dispersion, these nascent proteins linger long enough to regulate cascaded reactions on the same DNA. We rationally designed a pulsatile genetic circuit by encoding an activator and repressor in feedback on the same DNA molecule. Driven by the local synthesis of only several proteins per hour and gene, the circuit dynamics exhibited enhanced variability between individual DNA molecules, and fluctuations with a broad power spectrum. Our results demonstrate that coexpressional localization, as a nonequilibrium process, facilitates single-DNA genetic circuits as dissipative nanodevices, with implications for nanobiotechnology applications and artificial cell design.

synthetic biology↗

bioRxiv2020

Decision-making in a synthetic cell: the limits of biological computation

We measured the dynamics of decision-making by a minimal bistable gene network integrated in a synthetic cell model, free of external perturbations. Reducing the number of gene copies from 105 to about 10 per cell revealed a transition from deterministic and slow computation to a fuzzy and rapid regime dominated by singleprotein fluctuations. Fuzzy computation appeared at DNA and protein concentrations 100-fold lower than necessary in equilibrium, suggesting rate enhancement by co-expressional localization. Whereas the high-copy regime was characterized by a sharp transition, hysteresis and robust memory, the low-copy limit showed incipient strong fluctuations, switching between states, and a signature of cellular individuality across the decision-making point. Our work establishes synthetic cells operating rapidly at the single molecule level to integrate gene regulatory networks with metabolic pathways for sustained survival with low energetic cost.One Sentence Summary Decision-making in a synthetic cell can be slow and precise or rapid and probabilistic by reducing the number of computing molecules by five decades down to single-molecule fluctuations.Competing Interest StatementThe Noireaux laboratory receives research funds from Arbor Biosciences, a distributor of the myTXTL cell-free protein synthesis kit.View Full Text

synthetic biology↗

bioRxiv2016

Control of Lipid Domain Organization by a Biomimetic Contractile Actomyosin Cortex

The cell membrane is a heterogeneously organized composite with lipid-protein micro-domains. The contractile actin cortex may govern the lateral organization of these domains in the cell membrane, yet the underlying mechanisms are not known. We recently reconstituted minimal actin cortices (MACs) (Vogel et al, 2013b) and here advanced our assay to investigate effects of rearranging actin filaments on the lateral membrane organization by introducing various phase-separated lipid mono-and bilayers to the MACs. The addition of actin filaments reorganized membrane domains. We found that the process reached a steady state where line tension and lateral crowding balanced. Moreover, the phase boundary allowed myosin driven actin filament rearrangements to actively move individual lipid domains, often accompanied by their shape change, fusion or splitting. Our findings illustrate how actin cortex remodeling in cells may control dynamic rearrangements of lipids and other molecules inside domains without directly binding to actin filaments.

synthetic biology↗