Quantifying E. coli proteome and transcriptome with single-molecule sensitivity in single cells.
Yuichi Taniguchi,Paul J. Choi,Gene-Wei Li,Huiyi Chen,Mohan Babu,Jeremy Hearn,Andrew Emili,X. Sunney Xie +7 more
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TLDR
System-wide analyses of protein and mRNA expression in individual cells with single-molecule sensitivity using a newly constructed yellow fluorescent protein fusion library for Escherichia coli found that almost all protein number distributions can be described by the gamma distribution with two fitting parameters which, at low expression levels, have clear physical interpretations as the transcription rate and protein burst size.Abstract:
Protein and messenger RNA (mRNA) copy numbers vary from cell to cell in isogenic bacterial populations. However, these molecules often exist in low copy numbers and are difficult to detect in single cells. We carried out quantitative system-wide analyses of protein and mRNA expression in individual cells with single-molecule sensitivity using a newly constructed yellow fluorescent protein fusion library for Escherichia coli. We found that almost all protein number distributions can be described by the gamma distribution with two fitting parameters which, at low expression levels, have clear physical interpretations as the transcription rate and protein burst size. At high expression levels, the distributions are dominated by extrinsic noise. We found that a single cell's protein and mRNA copy numbers for any given gene are uncorrelated.read more
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Quantitative influence of macromolecular crowding on gene regulation kinetics.
TL;DR: The model compares the model to experimental data for LacI repressor and finds that non-specific binding of the protein to DNA is activation- and not diffusion-limited, and shows that optimal searching strategy depends on TF abundance.
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Precision of Readout at the hunchback Gene: Analyzing Short Transcription Time Traces in Living Fly Embryos
Jonathan Desponds,Jonathan Desponds,Jonathan Desponds,Huy Tran,Huy Tran,Huy Tran,Teresa Ferraro,Teresa Ferraro,Teresa Ferraro,Tanguy Lucas,Tanguy Lucas,Tanguy Lucas,Carmina Angelica Perez Romero,Aurelien Guillou,Aurelien Guillou,Aurelien Guillou,Cécile Fradin,Mathieu Coppey,Mathieu Coppey,Mathieu Coppey,Nathalie Dostatni,Nathalie Dostatni,Nathalie Dostatni,Aleksandra M. Walczak,Aleksandra M. Walczak,Aleksandra M. Walczak +25 more
TL;DR: It is shown that the precision of the expression of the hunchback gene to measure its position along the anterior-posterior axis is low both at the boundary and in the anterior even at cycle 13, suggesting additional post-transcriptional averaging mechanisms to provide the precision observed in fixed embryos.
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Millisecond single-molecule localization microscopy combined with convolution analysis and automated image segmentation to determine protein concentrations in complexly structured, functional cells, one cell at a time
Adam J. M. Wollman,Mark C. Leake +1 more
TL;DR: CoPro as mentioned in this paper uses millisecond imaging with convolution analysis, automated image segmentation and super-resolution localization microscopy to generate robust estimates for protein concentration in different compartments of single living cells, validated using realistic simulations of complex multiple compartment cell types.
Journal ArticleDOI
Connecting protein and mRNA burst distributions for stochastic models of gene expression
TL;DR: In this paper, the authors derived analytical expressions connecting protein and mRNA burst distributions, which showed how the functional form of the mRNA burst distribution can be inferred from the protein burst distribution.
Journal ArticleDOI
Rickettsia typhi possesses phospholipase A2 enzymes that are involved in infection of host cells.
M. Sayeedur Rahman,Joseph J. Gillespie,Joseph J. Gillespie,Simran J. Kaur,Khandra T. Sears,Shane M. Ceraul,Magda Beier-Sexton,Abdu F. Azad +7 more
TL;DR: This study suggests that R. typhi utilizes two evolutionary divergent patatin phospholipases to support its intracellular life cycle, a mechanism distinguishing it from other rickettsial species.
References
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Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection.
Tomoya Baba,Takeshi Ara,Miki Hasegawa,Yuki Takai,Yoshiko Okumura,Miki Baba,Kirill A. Datsenko,Masaru Tomita,Barry L. Wanner,Hirotada Mori,Hirotada Mori +10 more
TL;DR: These mutants—the ‘Keio collection’—provide a new resource not only for systematic analyses of unknown gene functions and gene regulatory networks but also for genome‐wide testing of mutational effects in a common strain background, E. coli K‐12 BW25113.
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Stochastic Gene Expression in a Single Cell
TL;DR: This work constructed strains of Escherichia coli that enable detection of noise and discrimination between the two mechanisms by which it is generated and reveals how low intracellular copy numbers of molecules can fundamentally limit the precision of gene regulation.
Journal ArticleDOI
Global analysis of protein localization in budding yeast
Won-Ki Huh,James V. Falvo,Luke C. Gerke,Adam S. Carroll,Russell W. Howson,Jonathan S. Weissman,Erin K. O'Shea +6 more
TL;DR: The construction and analysis of a collection of yeast strains expressing full-length, chromosomally tagged green fluorescent protein fusion proteins helps reveal the logic of transcriptional co-regulation, and provides a comprehensive view of interactions within and between organelles in eukaryotic cells.
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Fabrication of microfluidic systems in poly(dimethylsiloxane)
J.C. McDonald,David C. Duffy,Janelle R. Anderson,Daniel T. Chiu,Hongkai Wu,Olivier Schueller,George M. Whitesides +6 more
TL;DR: Fabrication of microfluidic devices in poly(dimethylsiloxane) (PDMS) by soft lithography provides faster, less expensive routes to devices that handle aqueous solutions.