Teahan, J. | Search bioRxiv

bioRxiv2020

Scanning ion conductance microscopy reveals differences in the ionic environments of gram positive and negative bacteria

This paper reports on the use of scanning ion conductance microscopy (SICM) to locally map the ionic properties and charge environment of two live bacterial strains: the gramnegative Escherichia coli and the gram-positive Bacillus subtilis. SICM results find heterogeneities across the bacterial surface, and significant differences among the grampositive and -negative bacteria. The bioelectrical environment of the B. subtilis was found to be considerably more negatively charged compared to E. coli. SICM measurements, fitted to a simplified finite element method (FEM) model, revealed surface charge values of -80 to -140 mC m-2 for the gram-negative E. coli. The gram-positive B. subtilis show a much higher conductivity around the cell wall, and surface charge values between -350 and -450 mC m-2 were found using the same simplified model. SICM was also able to detect regions of high negative charge near B. subtilis, not detected in the topographical SICM response and attributed to extracellular polymeric substance. To further explore how the B. subtilis cell wall structure can influence the SICM current response, a more comprehensive FEM model, accounting for the physical properties of the gram-positive cell wall, was developed. The new model provides a more realistic description of the cell wall and allowed investigation of the relation between its key properties and SICM currents, building foundations to further investigate and improve understanding of the gram-positive cellular microenvironment. O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

microbiology↗

bioRxiv2019

Polymerase recycling contributes to transcriptional noise

BackgroundTranscription in mammalian cells is a complex stochastic process involving shuttling of polymerase between genes and phase-separated liquid condensates. It occurs in bursts, which results in vastly different numbers of an mRNA species in isogenic cell populations. Several factors contributing to transcriptional bursting have been identified, usually classified as intrinsic, in other words local to single genes, or extrinsic, relating to the macroscopic state of the cell. However, some possible contributors have not been explored yet. Here, we focus on processes at the 3 and 5 ends of a gene that enable reinitiation of transcription upon termination. ResultsUsing Bayesian methodology, we measure the transcriptional bursting in inducible transgenes, showing that perturbation of polymerase shuttling typically reduces burst size, increases burst frequency, and thus limits transcriptional noise. Analysis based on paired-end tag sequencing (PolII ChIA-PET) suggests that this effect is genome wide. The observed noise patterns are also reproduced by a generative model that captures major characteristics of the polymerase flux between the ends of a gene and a phase-separated compartment. ConclusionsInteractions between the 3 and 5 ends of a gene, which facilitate polymerase recycling, are major contributors to transcriptional noise.

systems biology↗