Enterohemorrhagic Escherichia coli infection inhibits colonic thiamin pyrophosphate uptake via transcriptional mechanism.

Kasin Yadunandam Anandam,Subrata Sabui,Sreya Subramanian,Morgan M Thompson,Hamid M Said

doi:10.1371/journal.pone.0224234

Kasin Yadunandam Anandam, Subrata Sabui + Show 3 more

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https://doi.org/10.1371/journal.pone.0224234

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Abstract

Colonocytes possess a specific carrier-mediated uptake process for the microbiota-generated thiamin (vitamin B1) pyrophosphate (TPP) that involves the TPP transporter (TPPT; product of the SLC44A4 gene). Little is known about the effect of exogenous factors (including enteric pathogens) on the colonic TPP uptake process. Our aim in this study was to investigate the effect of Enterohemorrhagic Escherichia coli (EHEC) infection on colonic uptake of TPP. We used human-derived colonic epithelial NCM460 cells and mice in our investigation. The results showed that infecting NCM460 cells with live EHEC (but not with heat-killed EHEC, EHEC culture supernatant, or with non-pathogenic E. Coli) to lead to a significant inhibition in carrier-mediated TPP uptake, as well as in level of expression of the TPPT protein and mRNA. Similarly, infecting mice with EHEC led to a significant inhibition in colonic TPP uptake and in level of expression of TPPT protein and mRNA. The inhibitory effect of EHEC on TPP uptake by NCM460 was found to be associated with reduction in the rate of transcription of the SLC44A4 gene as indicated by the significant reduction in the activity of the SLC44A4 promoter transfected into EHEC infected cells. The latter was also associated with a marked reduction in the level of expression of the transcription factors CREB-1 and ELF3, which are known to drive the activity of the SLC44A4 promoter. Finally, blocking the ERK1/2 and NF-kB signaling pathways in NCM460 cells significantly reversed the level of EHEC inhibition in TPP uptake and TPPT expression. Collectively, these findings show, for the first time, that EHEC infection significantly inhibit colonic uptake of TPP, and that this effect appears to be exerted at the level of SLC44A4 transcription and involves the ERK1/2 and NF-kB signaling pathways.

Highlights

Thiamin pyrophosphate (TPP; called thiamin di-phosphate), is a biologically active form of vitamin B1 that acts as a cofactor for multiple enzymes that are involved in critical metabolic reactions (e. g., energy metabolism, reduction of cellular oxidative stress) [1, 2, 3]
Our results showed that Enterohemorrhagic E. coli (EHEC) infection causes a significant inhibition in colonic TPP uptake and that this inhibition is exerted at the level of transcription of the SLC44A4 gene and involves the ERK1/2 and NF-kB signaling pathways
We examined the effect of EHEC infection, as well as that of the non-pathogenic E. coli HS4 (200 multiplicity of infection (MOI)), of colonic epithelial NCM460 cells on TPP uptake

Summary

Introduction

Thiamin pyrophosphate (TPP; called thiamin di-phosphate), is a biologically active form of vitamin B1 that acts as a cofactor for multiple enzymes (pyruvate dehydrogenase, α-ketoglutarate dehydrogenase, branched-chain α-ketoacid dehydrogenase, transketolase) that are involved in critical metabolic reactions (e. g., energy metabolism, reduction of cellular oxidative stress) [1, 2, 3]. Studies from our laboratory have shown that the human/mammalian colonocytes are capable of absorbing both of these forms of the vitamin and that this occurs via distinct and specific carrier-mediated mechanisms [19,20,21]. Large intestinal absorption of free thiamin involves the SLC19A2 and SLC19A3 uptake systems [12, 19, 22], while that of TPP involves the recently identified TPPT system (product of the SLC44A4 gene) [20, 21]. Our results showed that EHEC infection causes a significant inhibition in colonic TPP uptake and that this inhibition is exerted at the level of transcription of the SLC44A4 gene and involves the ERK1/2 and NF-kB signaling pathways

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