Synthetic biology approach for the design of butyrate producing bacteria for colon cancer therapy

Kızıl, Cihat Burak
Butyric acid is a short chain fatty acid (SCFA) that is generated in the colon by butyrate-producing bacteria. Butyrate, along with other SCFAs, serve as substrates for energy production in the gut epithelium. Additionally, these compounds are known to reduce inflammation and enhance differentiation of mammalian cells. Several species of bacteria are known to produce butyrate in the gut through different biochemical pathways. In this study we have aimed to generate a butyrate producing commensal E. coli K12 strain that is known to be non butyrogenic. For this, we have used a strain in which fermentation related pathways have been deleted (ΔadhE, Δldha, ΔackA-pta, ΔfrdC) while enzymes for the reversal of β-oxidation (AtoB, FadB, FabI) and butyrate production [Butyryl-CoA: Acetate-CoA transferase (ButCoAT)] have been overexpressed. The E. coli K12 strain engineered with these alterations was capable of synthesizing mM amounts of butyrate. Caco-2 colon cancer cell line was incubated with a 1:1(v/v) mixture of conditioned medium generated from the engineered E. coli K12 strain and culture medium. A strong activation of the Mitogen Activated Protein Kinase (MAPK) family protein p38 and its downstream target MAPKAPK2 (MK2) was observed in cells treated with the conditioned medium compared to controls. The MAPK pathway orchestrates several regulatory mechanisms in epithelial cells. Our study suggests that engineered bacteria can be used to regulate gene expression in the gut.


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Sodium butyrate (NaBt) is a four-carbon short chain fatty acid, produced naturally in colon as the end product of the bacterial anaerobic metabolism on dietary fibers. It was previously shown that NaBt can induce differentiation and may inhibit proliferation. The objective of this study was to investigate the effect of NaBt-induced differentation on inflammatory pathways in HT29 colon cancer cells. For this purpose, first, cells were treated with varying concentrations of NaBt from 1-5 mM and amount require...
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Ülgen, Doğukan Hazar; Banerjee, Sreeparna; Department of Biology (2015)
Sodium butyrate (NaBt) is a four-carbon short-chain fatty acid histone deacetylase inhibitor (HDACi) that is available in the colon through the commensal microbiota-mediated fermentation of dietary fibers. It is the main source of energy for colonocytes, and is regarded to have tumor suppressive effects, most prominently in colorectal cancer (CRC). Cyclooxygenase-2 (COX-2) is a gene important in the inflammatory response due to its ability to convert arachidonic acid to prostaglandins. Overexpression and ov...
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The conversion of the cellular form of the prion protein (PrPC) to an abnormal, alternatively folded isoform (PrPSc) is the central event in prion diseases or transmissible spongiform encephalopathies. Recent studies have demonstrated de novo generation of murine prions from recombinant prion protein (recPrP) after inoculation into transgenic and wild-type mice. These so-called synthetic prions lead to novel prion diseases with unique neuropathological and biochemical features. Moreover, the use of recPrP i...
Production of an acetone-butanol-ethanol mixture from Clostridium acetobutylicum and its conversion to high-value biofuels
Sreekumar, Sanil; Baer, Zachary C.; Pazhamalai, Anbarasan; Günbaş, Emrullah Görkem; Grippo, Adam; Blanch, Harvey W.; Clark, Douglas S.; Toste, F. Dean (Springer Science and Business Media LLC, 2015-03-01)
Clostridium acetobutylicum is a bacterial species that ferments sugar to a mixture of organic solvents (acetone, butanol and ethanol). This protocol delineates a methodology to combine solventogenic clostridial fermentation and chemical catalysis via extractive fermentation for the production of biofuel blendstocks. Extractive fermentation of C. acetobutylicum is operated in fed-batch mode with a concentrated feed solution (500 grams per liter glucose and 50 grams per liter yeast extract) for 60 h, producin...
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Cingöz, Harun; Erson Bensan, Ayşe Elif; Department of Biology (2019)
Sorting Nexin 3 (SNX3) is part of the retromer complex that recycles cargo receptors back to plasma membrane or to Trans Golgi Network. WNT ligand carrier protein Wntless (WLS) is a known SNX3 cargo protein. Our earlier data suggested over expression of SNX3 in colon cancer cells. Considering its importance in receptor recycling, we hypothesized SNX3 to be a potential modulator of cancer related receptors. To begin understanding the role of SNX3, we developed RNAi models of SNX3 in SW480 colon cancer cells ...
Citation Formats
C. B. Kızıl, “Synthetic biology approach for the design of butyrate producing bacteria for colon cancer therapy,” Thesis (M.S.) -- Graduate School of Natural and Applied Sciences. Biology., Middle East Technical University, 2019.