genetic-engineering-dictyostelium-discoideum-cells-based-on-selection

Genetic Engineering of <em>Dictyostelium discoideum</em> Cells Based on Selection and Growth on Bacteria

Dictyostelium discoideum is an intriguing model organism for the study of cell differentiation processes during development, cell signaling, and other ...

University College London

Transcriptional regulation is noisy, yet despite this variability, embryonic development reproducibly generates form and function. Recent work demonstrates that patterns of transcriptional activity in embryos are stably inherited through mitosis. These observations have implications for how accuracy arises in development. 

Gene Regulation: Stable Noise.

Gene expression levels vary greatly within similar cells, even within clonal cell populations [1]. These spontaneous expression differences underlie cell fate diversity in both differentiation and disease [2]. The mechanisms responsible for generating expression variability are poorly understood. Using single-cell transcriptomics, we show that transcript variability emerging during Dictyostelium differentiation is driven predominantly by repression rather than activation. The increased variability of repressed genes was observed over a broad range of expression levels, indicating that variability is actively imposed and not a passive statistical effect of the reduced numbers of molecules accompanying repression. These findings can be explained by a simple model of transcript production, with expression controlled by the frequency, rather than the magnitude, of transcriptional firing events. Our study reveals that the generation of differences between cells can be a direct consequence of the basic mechanisms of transcriptional regulation.

Generation of Single-Cell Transcript Variability by Repression.

During the evolution of gene families, functional diversification of proteins often follows gene duplication. However, many gene families expand while preserving protein sequence. Why do cells maintain multiple copies of the same gene? Here we have addressed this question for an actin family with 17 genes encoding an identical protein. The genes have divergent flanking regions and are scattered throughout the genome. Surprisingly, almost the entire family showed similar developmental expression profiles, with their expression also strongly coupled in single cells. Using live cell imaging, we show that differences in gene expression were apparent over shorter timescales, with family members displaying different transcriptional bursting dynamics. Strong "bursty" behaviors contrasted steady, more continuous activity, indicating different regulatory inputs to individual actin genes. To determine the sources of these different dynamic behaviors, we reciprocally exchanged the upstream regulatory regions of gene family members. This revealed that dynamic transcriptional behavior is directly instructed by upstream sequence, rather than features specific to genomic context. A residual minor contribution of genomic context modulates the gene OFF rate. Our data suggest promoter diversification following gene duplication could expand the range of stimuli that regulate the expression of essential genes. These observations contextualize the significance of transcriptional bursting.

Article	Year
Gene Regulation: Stable Noise. Current biology : CB\| PubMed ID: 26811888	2016
Generation of Single-Cell Transcript Variability by Repression. Current biology : CB\| PubMed ID: 28602650	2017
Promoter-mediated diversification of transcriptional bursting dynamics following gene duplication. Proceedings of the National Academy of Sciences of the United States of America\| PubMed ID: 30061408	2018

Jonathan R. Chubb

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