12 research outputs found
Protein Pattern Formation
Protein pattern formation is essential for the spatial organization of many
intracellular processes like cell division, flagellum positioning, and
chemotaxis. A prominent example of intracellular patterns are the oscillatory
pole-to-pole oscillations of Min proteins in \textit{E. coli} whose biological
function is to ensure precise cell division. Cell polarization, a prerequisite
for processes such as stem cell differentiation and cell polarity in yeast, is
also mediated by a diffusion-reaction process. More generally, these functional
modules of cells serve as model systems for self-organization, one of the core
principles of life. Under which conditions spatio-temporal patterns emerge, and
how these patterns are regulated by biochemical and geometrical factors are
major aspects of current research. Here we review recent theoretical and
experimental advances in the field of intracellular pattern formation, focusing
on general design principles and fundamental physical mechanisms.Comment: 17 pages, 14 figures, review articl
Efficient switching of mCherry fluorescence using chemical caging
Fluorophores with dynamic or controllable fluorescence emission have become essential tools for advanced imaging, such as superresolution imaging. These applications have driven the continuing development of photoactivatable or photoconvertible labels, including genetically encoded fluorescent proteins. These new probes work well but require the introduction of new labels that may interfere with the proper functioning of existing constructs and therefore require extensive functional characterization. In this work we show that the widely used red fluorescent protein mCherry can be brought to a purely chemically induced blue-fluorescent state by incubation with β-mercaptoethanol (βME). The molecules can be recovered to the red fluorescent state by washing out the βME or through irradiation with violet light, with up to 80% total recovery. We show that this can be used to perform single-molecule localization microscopy (SMLM) on cells expressing mCherry, which renders this approach applicable to a very wide range of existing constructs. We performed a detailed investigation of the mechanism underlying these dynamics, using X-ray crystallography, NMR spectroscopy, and ab initio quantum-mechanical calculations. We find that the βME-induced fluorescence quenching of mCherry occurs both via the direct addition of βME to the chromophore and through βME-mediated reduction of the chromophore. These results not only offer a strategy to expand SMLM imaging to a broad range of available biological models, but also present unique insights into the chemistry and functioning of a highly important class of fluorophores
Physical Views on ParABS-Mediated DNA Segregation
International audienceIn this chapter, we will focus on ParABS: an apparently simple, three-component system, required for the segregation of bacterial chromosomes and plasmids. We will specifically describe how biophysical measurements combined with physical modeling advanced our understanding of the mechanism of ParABS-mediated complex assembly, segregation and positioning