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

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.

Role of Dietary Micronutrients on Gut Microbial Dysbiosis and Modulation in Inflammatory Bowel Disease

“…[ 33 ] This colonic B1 absorption was shown to be inhibited during enterohemorrhagic Escherichia coli infection. [ 34 ] Interestingly, the gene responsible for colonic B1 absorption (SLC44A4) was recently identified as a UC susceptibility gene in an Indian cohort suggesting a potential role of colonic B1 uptake in UC pathology. [ 35 ] Although further studies investigating the interaction between B1‐microbiota‐host in IBD are necessary, increasing B1 levels through diet and gut bacteria may be a prominent strategy to induce gut homeostasis in IBD.…”

Hypoxia inhibits colonic uptake of the microbiota-generated forms of vitamin B1 via HIF-1α-mediated transcriptional regulation of their transporters

“…The proteins were then blotted onto polyvinylidene difluoride membranes and probed with anti-cTPPT (1:500), anti-THTR-1 (1:1000), anti-THTR-2 (1:1000), anti-HIF-1α (1:500), or anti-HIF-2α (1:500) antibodies and simultaneously with anti-GAPDH (1:2000) primary antibodies. Specificity of the anti-cTPPT, THTR-1, and THTR-2 antibodies was validated in our laboratory previously using different approaches that include overexpression of tag protein or gene silencing ( 52 , 53 ). Other antibodies were validated by either the respective company or by the other investigators using knockout animals and/or gene knockdown approaches.…”