Automated live-cell single-molecule tracking in enteroid monolayers reveals transcription factor dynamics probing lineage-determining function.

Lineage transcription factors (TFs) provide one regulatory level of differentiation crucial for the generation and maintenance of healthy tissues. To probe TF function by measuring their dynamics during adult intestinal homeostasis, we established HILO-illumination-based live-cell single-molecule tracking (SMT) in mouse small intestinal enteroid monolayers recapitulating tissue differentiation hierarchies in vitro. To increase the throughput, capture cellular features, and correlate morphological characteristics with diffusion parameters, we developed an automated imaging and analysis pipeline, broadly applicable to two-dimensional culture systems.

Thermodynamic architecture and conformational plasticity of GPCRs.

G-protein-coupled receptors (GPCRs) are ubiquitous integral membrane proteins involved in diverse cellular signaling processes. Here, we carry out a large-scale ensemble thermodynamic study of 45 ligand-free GPCRs employing a structure-based statistical mechanical framework. We find that multiple partially structured states co-exist in the GPCR native ensemble, with the TM helices 1, 6 and 7 displaying varied folding status, and shaping the conformational landscape.

Probing patterning in microbial consortia with a cellular automaton for spatial organisation.

Microbial consortia exhibit spatial patterning across diverse environments. Since probing the self-organization of natural microbial communities is limited by their inherent complexity, synthetic models have emerged as attractive alternatives. In this study, we develop novel frameworks of bacterial communication and explore the emergent spatiotemporal organization of microbes.

pPerturb: A Server for Predicting Long-Distance Energetic Couplings and Mutation-Induced Stability Changes in Proteins via Perturbations.

The strength of intraprotein interactions or contact network is one of the dominant factors determining the thermodynamic stabilities of proteins. The nature and the extent of connectivity of this network also play a role in allosteric signal propagation characteristics upon ligand binding to a protein domain. Here, we develop a server for rapid quantification of the strength of an interaction network by employing an experimentally consistent perturbation approach previously validated against a large data set of 375 mutations in 19 different proteins.