Transcriptional responses of human intestinal epithelial HT-29 cells to spore-displayed p40 derived from Lacticaseibacillus rhamnosus GG.

BACKGROUNDS: The aims of this study were to construct spore-displayed p40, a Lacticaseibacillus rhamnosus GG-derived soluble protein, using spore surface display technology and to evaluate transcriptional responses in human intestinal epithelial cells. RESULTS: p40 was displayed on the surface of Bacillus subtilis spores using spore coat protein CotG as an anchor protein. Effects of spore-displayed p40 (CotG-p40) on gene expression of intestinal epithelial cell line HT-29 were evaluated by transcriptome analysis using RNA-sequencing. As a result of differentially expressed gene (DEG) analysis, 81 genes were up-regulated and 82 genes were down-regulated in CotG-p40 stimulated cells than in unstimulated cells. Gene ontology enrichment analysis showed that CotG-p40 affected biological processes such as developmental process, metabolic process, cell surface receptor linked signaling pathway, and retinoic acid metabolic process. Gene-gene network analysis suggested that 10 DEGs (EREG, FOXF1, GLI2, PTGS2, SPP1, MMP19, TNFRSF1B, PTGER4, CLDN18, and ALDH1A3) activated by CotG-p40 were associated with probiotic action. CONCLUSIONS: This study demonstrates the regulatory effects of CotG-p40 on proliferation and homeostasis of HT-29 cells. This study provided comprehensive insights into the transcriptional response of human intestinal epithelial cells stimulated by CotG-p40.

Effects of Spore-Displayed p75 Protein from Lacticaseibacillus rhamnosus GG on the Transcriptional Response of HT-29 Cells.

A Lacticaseibacillus rhamnosus GG-derived protein, p75, is one of the key molecules exhibiting probiotic activity. However, the molecular mechanism and transcriptional response of p75 in human intestinal epithelial cells are not completely understood. To gain a deeper understanding of its potential probiotic action, this study investigated genome-wide responses of HT-29 cells to stimulation by spore-displayed p75 (CotG-p75) through a transcriptome analysis based on RNA sequencing. Analysis of RNA-seq data showed significant changes of gene expression in HT-29 cells stimulated by CotG-p75 compared to the control. A total of 189 up-regulated and 314 down-regulated genes was found as differentially expressed genes. Gene ontology enrichment analysis revealed that a large number of activated genes was involved in biological processes, such as epithelial cell differentiation, development, and regulation of cell proliferation. A gene-gene interaction network analysis showed that several DEGs, including AREG, EREG, HBEGF, EPGN, FASLG, GLI2, CDKN1A, FOSL1, MYC, SERPINE1, TNFSF10, BCL6, FLG, IVL, SPRR1A, SPRR1B, SPRR3, and MUC5AC, might play a critical role in these biological processes. RNA-seq results for selected genes were verified by reverse transcription-quantitative polymerase chain reaction. Overall, these results provide extensive knowledge about the transcriptional responses of HT-29 cells to stimulation by CotG-p75. This study showed that CotG-p75 can contribute to cell survival and epithelial development in human intestinal epithelial cells.