A Synthetic Biology Approach to Sequential Stripe Patterning and Somitogenesis

AbstractReaction-diffusion (RD) based clock and wavefront model has long been proposed as the mechanism underlying biological pattern formation of repeated and segmented structures including somitogenesis. However, systematic molecular level understanding of the mechanism remains elusive, largely due to the lack of suitable experimental systems to probe RD quantitatively in vivo. Here we design a synthetic gene circuit that couples gene expression regulation (reaction) with quorum sensing (diffusion) to guide bacterial cells self-organizing into stripe patterns at both microscopic and colony scales. An experimentally verified mathematical model confirms that these periodic spatial structures are emerged from the integration of oscillatory gene expression as the molecular clock and the outward expanding diffusions as the propagating wavefront. Furthermore, our paired model-experiment data illustrate that the RD-based patterning is sensitive to initial conditions and can be modulated by external inducers to generate diverse patterns, including multiple-stripe pattern, target-like pattern and ring patterns with reversed fluorescence. Powered by our synthetic biology setup, we also test different topologies of gene networks and show that network motifs enabling robust oscillations are foundations of sequential stripe pattern formation. These results verified close connections between gene network topology and resulting RD driven pattern formation, offering an engineering approach to help understand biological development.

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Gerea: Prediction Of Gene Expression Regulators From Transcriptome Profiling Data To Transition Networks

Current Bioinformatics ◽

10.2174/1574893616666210621100335 ◽

2021 ◽

Vol 16 ◽

Author(s):

Min Yao ◽

Caiyun Jiang ◽

Chenglong Li ◽

Yongxia Li ◽

Shan Jiang ◽

...

Keyword(s):

Gene Expression ◽

Gene Networks ◽

Target Gene ◽

Gene Expression Regulation ◽

Transcriptome Profiling ◽

Expression Regulation ◽

Mouse Gene ◽

Mouse Gene Expression ◽

Causality Inference ◽

Identify Gene Expression

Background: Mammalian genes are regulated at the transcriptional and post-transcriptional levels. These mechanisms may involve the direct promotion or inhibition of transcription via a regulator or post-transcriptional regulation through factors such as micro (mi)RNAs. Objective: This study aimed to construct gene regulation relationships modulated by causality inference-based miRNA-(transition factor)-(target gene) networks and analyze gene expression data to identify gene expression regulators. Methods: Mouse gene expression regulation relationships were manually curated from literature using a text mining method which was then employed to generate miRNA-(transition factor)-(target gene) networks. An algorithm was then introduced to identify gene expression regulators from transcriptome profiling data by applying enrichment analysis to these networks. Results: A total of 22,271 mouse gene expression regulation relationships were curated for 4,018 genes and 242 miRNAs. GEREA software was developed to perform the integrated analyses. We applied the algorithm to transcriptome data for synthetic miR-155 oligo-treated mouse CD4+ T-cells and confirmed that miR-155 is an important network regulator. The software was also tested on publicly available transcriptional profiling data for Salmonella infection, resulting in the identification of miR-125b as an important regulator. Conclusion: The causality inference-based miRNA-(transition factor)-(target gene) networks serve as a novel resource for gene expression regulation research, and GEREA is an effective and useful adjunct to the currently available methods. The regulatory networks and the algorithm implemented in the GEREA software package are available under a free academic license at website : http://www.thua45.cn/gerea.

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Comparison of gene expression and activation of transcription factors in organoid-derived monolayer intestinal epithelial cells and organoids

Bioscience Biotechnology and Biochemistry ◽

10.1093/bbb/zbab136 ◽

2021 ◽

Author(s):

Yu Takahashi ◽

Yu Inoue ◽

Keitaro Kuze ◽

Shintaro Sato ◽

Makoto Shimizu ◽

...

Keyword(s):

Gene Expression ◽

Epithelial Cells ◽

Gene Expression Regulation ◽

Intestinal Epithelial Cells ◽

Three Dimensional ◽

Culture Conditions ◽

Dependent Manner ◽

Intestinal Epithelial

Abstract Intestinal organoids better represent in vivo intestinal properties than conventionally used established cell lines in vitro. However, they are maintained in three-dimensional culture conditions that may be accompanied by handling complexities. We characterized the properties of human organoid-derived two-dimensionally cultured intestinal epithelial cells (IECs) compared with those of their parental organoids. We found that the expression of several intestinal markers and functional genes were indistinguishable between monolayer IECs and organoids. We further confirmed that their specific ligands equally activate intestinal ligand-activated transcriptional regulators in a dose-dependent manner. The results suggest that culture conditions do not significantly influence the fundamental properties of monolayer IECs originating from organoids, at least from the perspective of gene expression regulation. This will enable their use as novel biological tools to investigate the physiological functions of the human intestine.

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MicroRNAs in mouse models of lymphoid malignancies

Journal of Nucleic Acids Investigation ◽

10.4081/jnai.2010.1727 ◽

2010 ◽

Vol 1 (1) ◽

pp. 8 ◽

Cited By ~ 5

Author(s):

Nicola A. O. Zanesi ◽

Yuri Pekarsky ◽

Francesco Trapasso ◽

George Calin ◽

Carlo M. Croce

Keyword(s):

Gene Expression ◽

Mouse Models ◽

Gene Expression Regulation ◽

Human Leukemia ◽

Down Regulation ◽

Over Expression ◽

Lymphoid Malignancies ◽

Knockout Animals ◽

Mouse Modeling

The discovery of microRNAs (miRNAs) has revealed a new layer of gene expression regulation that affects many normal and pathologic biological systems. Among the malignancies affected by the dysregulation of miRNAs there are cancers of lymphoid origin, in which miRNAs are thought to have tumor suppressive or tumor promoting activities, depending on the nature of their specific targets. In the last 4-5 years, the experimental field that provided the deepest insights into the in vivo biology of miRNAs is that of mouse modeling in which transgenic and knockout animals mimic, respectively, over-expression or down-regulation of specific miRNAs involved in human leukemia/lymphoma. This review discusses recent advances in our understanding of lymphoid malignancies based on the natural and engineered mouse models of three different miRNAs, miR-15a/16-1 cluster, miR-155, and miR-17-92 cluster.

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Synthetic protein-binding DNA sponge as a tool to tune gene expression and mitigate protein toxicity

Nature Communications ◽

10.1038/s41467-020-19552-9 ◽

2020 ◽

Vol 11 (1) ◽

Author(s):

Xinyi Wan ◽

Filipe Pinto ◽

Luyang Yu ◽

Baojun Wang

Keyword(s):

Gene Expression ◽

Gene Regulation ◽

Protein Binding ◽

Gene Networks ◽

Dynamic Range ◽

Gene Expression Regulation ◽

Single Layer ◽

Regulation Mechanism ◽

Gene Circuits ◽

Synthetic Dna

AbstractVersatile tools for gene expression regulation are vital for engineering gene networks of increasing scales and complexity with bespoke responses. Here, we investigate and repurpose a ubiquitous, indirect gene regulation mechanism from nature, which uses decoy protein-binding DNA sites, named DNA sponge, to modulate target gene expression in Escherichia coli. We show that synthetic DNA sponges can be designed to reshape the response profiles of gene circuits, lending multifaceted tuning capacities including reducing basal leakage by >20-fold, increasing system output amplitude by >130-fold and dynamic range by >70-fold, and mitigating host growth inhibition by >20%. Further, multi-layer DNA sponges for decoying multiple regulatory proteins provide an additive tuning effect on the responses of layered circuits compared to single-layer sponges. Our work shows synthetic DNA sponges offer a simple yet generalizable route to systematically engineer the performance of synthetic gene circuits, expanding the current toolkit for gene regulation with broad potential applications.

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Receptor-Based Models with Diffusion-Driven Instability for Pattern Formation in Hydra

Journal of Biological System ◽

10.1142/s0218339003000889 ◽

2003 ◽

Vol 11 (03) ◽

pp. 293-324 ◽

Cited By ~ 16

Author(s):

Anna Marciniak-Czochra

Keyword(s):

Pattern Formation ◽

Feedback Loop ◽

De Novo ◽

Initial Conditions ◽

Reaction Diffusion ◽

Reaction Diffusion Equations ◽

Variable Model ◽

Dissociated Cells ◽

Cutting Experiments ◽

Positional Value

The aim of this paper is to show under which conditions a receptor-based model can produce and regulate patterns. Such model is applied to the pattern formation and regulation in a fresh water polyp, hydra. The model is based on the idea that both head and foot formation could be controlled by receptor-ligand binding. Positional value is determined by the density of bound receptors. The model is defined in the form of reaction-diffusion equations coupled with ordinary differential equations. The objective is to check what minimal processes are sufficient to produce patterns in the framework of a diffusion-driven (Turing-type) instability. Three-variable (describing the dynamics of ligands, free and bound receptors) and four-variable models (including also an enzyme cleaving the ligand) are analyzed and compared. The minimal three-variable model takes into consideration the density of free receptors, bound receptors and ligands. In such model patterns can evolve only if self-enhancement of free receptors, i.e., a positive feedback loop between the production of new free receptors and their present density, is assumed. The final pattern strongly depends on initial conditions. In the four-variable model a diffusion-driven instability occurs without the assumption that free receptors stimulate their own synthesis. It is shown that gradient in the density of bound receptors occurs if there is also a second diffusible substance, an enzyme, which degrades ligands. Numerical simulations are done to illustrate the analysis. The four-variable model is able to capture some results from cutting experiments and reflects de novo pattern formation from dissociated cells.

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Regulating synthetic gene networks in 3D materials

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1204705109 ◽

2012 ◽

Vol 109 (38) ◽

pp. 15217-15222 ◽

Cited By ~ 28

Author(s):

Tara L. Deans ◽

Anirudha Singh ◽

Matthew Gibson ◽

Jennifer H. Elisseeff

Keyword(s):

Synthetic Biology ◽

Gene Networks ◽

Materials Science ◽

Genetic Circuits ◽

Therapeutic Applications ◽

Regulatory Processes ◽

Cellular Events ◽

The Matrix

Combining synthetic biology and materials science will enable more advanced studies of cellular regulatory processes, in addition to facilitating therapeutic applications of engineered gene networks. One approach is to couple genetic inducers into biomaterials, thereby generating 3D microenvironments that are capable of controlling intrinsic and extrinsic cellular events. Here, we have engineered biomaterials to present the genetic inducer, IPTG, with different modes of activating genetic circuits in vitro and in vivo. Gene circuits were activated in materials with IPTG embedded within the scaffold walls or chemically linked to the matrix. In addition, systemic applications of IPTG were used to induce genetic circuits in cells encapsulated into materials and implanted in vivo. The flexibility of modifying biomaterials with genetic inducers allows for patterned placement of these inducers that can be used to generate distinct patterns of gene expression. Together, these genetically interactive materials can be used to characterize genetic circuits in environments that more closely mimic cells’ natural 3D settings, to better explore complex cell–matrix and cell–cell interactions, and to facilitate therapeutic applications of synthetic biology.

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Inactivation of the Euchromatic Histone-Lysine N-Methyltransferase 2 Pathway in Pancreatic Epithelial Cells Antagonizes Cancer Initiation and Pancreatitis-Associated Promotion by Altering Growth and Immune Gene Expression Networks

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.681153 ◽

2021 ◽

Vol 9 ◽

Author(s):

Guillermo Urrutia ◽

Thiago Milech de Assuncao ◽

Angela J. Mathison ◽

Ann Salmonson ◽

Romica Kerketta ◽

...

Keyword(s):

Gene Expression ◽

Gene Networks ◽

Ex Vivo ◽

Mapk Pathway ◽

Intraepithelial Neoplasia ◽

Ductal Adenocarcinoma ◽

Immune Gene ◽

Protein Levels ◽

Histone Lysine

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive, painful disease with a 5-year survival rate of only 9%. Recent evidence indicates that distinct epigenomic landscapes underlie PDAC progression, identifying the H3K9me pathway as important to its pathobiology. Here, we delineate the role of Euchromatic Histone-lysine N-Methyltransferase 2 (EHMT2), the enzyme that generates H3K9me, as a downstream effector of oncogenic KRAS during PDAC initiation and pancreatitis-associated promotion. EHMT2 inactivation in pancreatic cells reduces H3K9me2 and antagonizes KrasG12D-mediated acinar-to-ductal metaplasia (ADM) and Pancreatic Intraepithelial Neoplasia (PanIN) formation in both the Pdx1-Cre and P48Cre/+KrasG12D mouse models. Ex vivo acinar explants also show impaired EGFR-KRAS-MAPK pathway-mediated ADM upon EHMT2 deletion. Notably, KrasG12D increases EHMT2 protein levels and EHMT2-EHMT1-WIZ complex formation. Transcriptome analysis reveals that EHMT2 inactivation upregulates a cell cycle inhibitory gene expression network that converges on the Cdkn1a/p21-Chek2 pathway. Congruently, pancreas tissue from KrasG12D animals with EHMT2 inactivation have increased P21 protein levels and enhanced senescence. Furthermore, loss of EHMT2 reduces inflammatory cell infiltration typically induced during KrasG12D-mediated initiation. The inhibitory effect on KrasG12D-induced growth is maintained in the pancreatitis-accelerated model, while simultaneously modifying immunoregulatory gene networks that also contribute to carcinogenesis. This study outlines the existence of a novel KRAS-EHMT2 pathway that is critical for mediating the growth-promoting and immunoregulatory effects of this oncogene in vivo, extending human observations to support a pathophysiological role for the H3K9me pathway in PDAC.

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Engineering pattern formation and morphogenesis

Biochemical Society Transactions ◽

10.1042/bst20200013 ◽

2020 ◽

Vol 48 (3) ◽

pp. 1177-1185

Author(s):

Jamie A. Davies ◽

Fokion Glykofrydis

Keyword(s):

Tissue Engineering ◽

Synthetic Biology ◽

Pattern Formation ◽

Research And Development ◽

Reaction Diffusion ◽

Future Research ◽

Self Organized ◽

Physical Form ◽

Basic Understanding ◽

Self Organizing

The development of natural tissues, organs and bodies depends on mechanisms of patterning and of morphogenesis, typically (but not invariably) in that order, and often several times at different final scales. Using synthetic biology to engineer patterning and morphogenesis will both enhance our basic understanding of how development works, and provide important technologies for advanced tissue engineering. Focusing on mammalian systems built to date, this review describes patterning systems, both contact-mediated and reaction-diffusion, and morphogenetic effectors. It also describes early attempts to connect the two to create self-organizing physical form. The review goes on to consider how these self-organized systems might be modified to increase the complexity and scale of the order they produce, and outlines some possible directions for future research and development.

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Spatio-temporal Brusselator Model and Biological Pattern Formation

Annual Research & Review in Biology ◽

10.9734/arrb/2021/v36i530380 ◽

2021 ◽

pp. 88-99

Author(s):

Zakir Hossine ◽

Oishi Khanam ◽

Md. Mashih Ibn Yasin Adan ◽

Md. Kamrujjaman

Keyword(s):

Pattern Formation ◽

Diffusion Model ◽

Initial Conditions ◽

Neumann Boundary ◽

Reaction Diffusion ◽

Two Dimensions ◽

Brusselator Model ◽

Reaction Diffusion Model ◽

Biological Pattern Formation ◽

Spatio Temporal

This paper explores a two-species non-homogeneous reaction-diffusion model for the study of pattern formation with the Brusselator model. We scrutinize the pattern formation with initial conditions and Neumann boundary conditions in a spatially heterogeneous environment. In the whole investigation, we assume the case for random diffusion strategy. The dynamics of model behaviors show that the nature of pattern formation with varying parameters and initial conditions thoroughly. The model also studies in the absence of diffusion terms. The theoretical and numerical observations explain pattern formation using the reaction-diffusion model in both one and two dimensions.

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