Biology subjects

Pawlak, M. R.

Publications and source records attributed to Pawlak, M. R..

5 recordsLinked to original sources

bioRxiv2025

A Generalizable Tension Sensor Platform for Mechanotherapeutic Discovery

Mechanical forces are critical regulators of cellular function, and their modulation represents a promising therapeutic strategy across a range of diseases, including cancer and fibrosis. DNA-based molecular tension sensors (MTSs) have emerged as powerful tools for detecting receptor-specific cellular forces but remain limited by susceptibility to nuclease degradation and constrained ligand compatibility. Here, we outline these barriers to broader adoption and demonstrate how integrating established stabilization strategies effectively mitigates nuclease sensitivity. In addition, we introduce an engineered protein that covalently couples ligands to DNA in a modular, receptor-agnostic manner. Together, these innovations enable robust, nearly universal deployment of DNA-based MTSs across diverse experimental contexts and target proteins. We apply this enhanced platform to profile the mechanobiological effects of force-modulating drugs in both immortalized and primary cell lines. Unlike indirect or context-limited methods, this approach delivers direct, quantitative readouts of drug-induced changes in mechanical force transmission, offering a scalable path toward personalized mechanotherapeutic screening.

biophysics↗

bioRxiv2023

Emergent functional behaviors of ribozymes in oxychlorine brines indicate Mars could host a unique niche for molecular evolution

Mars is a particularly attractive candidate among known astronomical objects to potentially host life. Results from space exploration missions have provided insights into Martian geochemistry indicating oxychlorine species, particularly perchlorate, are ubiquitous features of the Martian geochemical landscape. Perchlorate presents potential obstacles for known forms of life due to its toxicity. However, it can also provide potential benefits, such as producing brines by deliquescence, like those thought to exist on present-day Mars. Here we show perchlorate brines support folding and catalysis of functional RNAs, while inactivating representative protein enzymes. Additionally, we show perchlorate and other oxychlorine species enable new ribozyme functions, including homeostasis-like regulatory behavior, copying of structured RNAs, and the first ribozyme-catalyzed chlorination of organic molecules. We suggest nucleic acids are uniquely well-suited to hypersaline Martian environments. Furthermore, Martian near-or subsurface oxychlorine brines, and brines found in potential lifeforms, could provide a unique niche for biomolecular evolution.

biochemistry↗

bioRxiv2022

RAD-TGTs: Measurement of cellular tensions via flow cytometry and DNA sequencing enabled by force-dependent rupture and delivery of DNA tension probes

Mechanical force is a key driver of cellular processes and is dysregulated in many diseases. Measuring cellular tensions to elucidate mechanotransduction pathways typically involves high-resolution but low throughput imaging of surfaces and arduous experimental preparation of materials. We present here Rupture and Deliver DNA-duplex based molecular tension sensors-RAD-TGTs. RAD-TGTs consist of immobilized DNA duplexes conjugated to a ligand and indicator (fluorophore, barcode etc) which rupture in a force-dependent manner when cells are bound. Readout of rupture is performed in cells of interest using high throughput methods such as flow cytometry and leveraging covalent DNA-protein linking HUH-tags simplifies the preparation of the tension sensor to allow use of "off-the-shelf" oligos. We demonstrate that rupture and delivery is decreased by inhibitors of cytoskeletal dynamics and knockout of mechanosensing proteins. We also show that rupture and delivery correlates with ligand affinity. Excitingly, we demonstrate that rupture and delivery of barcoded DNA-duplexes can be quantified using DNA sequencing, propelling cellular force measurements into the -omics era.

biophysics↗

bioRxiv2022

Controlled exchange of protein and nucleic acid signals from and between synthetic minimal cells

Synthetic minimal cells are a class of small liposome bioreactors that have some, but not all functions of live cells. Here, we report a critical step towards the development of a bottom-up minimal cell: cellular export of functional protein and RNA products. We used cell penetrating peptide tags to translocate payloads across a synthetic cell vesicle membrane. We demonstrated efficient transport of active enzymes, and transport of nucleic acid payloads by RNA binding proteins. We investigated influence of a concentration gradient alongside other factors on the efficiency of the translocation, and we show a method to increase product accumulation in one location. We demonstrate the use of this technology to engineer molecular communication between different populations of synthetic cells, to exchange protein and nucleic acid signals. The synthetic minimal cell production and export of proteins or nucleic acids allows experimental designs that approach the complexity and relevancy of natural biological systems.

synthetic biology↗

bioRxiv2019

DNA transistors switched by the Hofmeister effect

Here we demonstrate that a DNA nanodevice can perform switchable electron transfer. The nanodevice is comprised of two strands, one of which can be selectively switched between a G-quadruplex and duplex or single-stranded conformations. In the G-quadruplex state, it binds the cofactor hemin, enabling peroxidase activity. This switching ability arises from our discovery that perchlorate, a chaotropic Hofmeister ion, selectively destabilizes duplex over G-quadruplex DNA. By varying perchlorate concentration, we show that the device can be switched between states that do and do not catalyze electron transfer catalysis. State switching can be achieved in three ways: thermally, by dilution, or by concentration. In each case, when operated in the presence of the cofactor hemin, the device catalyzes electron transfer in only the G-quadruplex state.

synthetic biology↗