Microbial regulation of intestinal epithelial gene expression

Abstract The intestinal epithelium serves essential functions of dietary nutrient absorption and defense against microbial infections through the coordinated regulation of gene expression. Intestinal microbiota significantly influence gene transcription in intestinal epithelial cells (IECs), including both induction and suppression of distinct gene

sets. We have shown that many microbially-suppressed genes are under control of the host transcription factor (TF) HNF4A and are involved in differentiation and metabolism of absorptive enterocytes. Microbially-induced genes are predicted to be downstream of TF pathways including IRF and NFκB and include genes involved in

proliferation, inflammation, and immune defense. What remains unknown is if these distinct microbiota-induced and microbiota-suppressed TF pathways communicate with each other and how they are balanced within IECs to maintain intestinal homeostasis. Our preliminary studies have revealed that these two opposing TF

pathways directly communicate within IECs to promote homeostasis by regulating responses to microbes. We find that HNF4A activity in mouse IECs is suppressed by microbiota and protects against microbiota-driven intestinal inflammation. Using single cell RNA sequencing in wild-type gnotobiotic mice, we have uncovered

significant heterogeneity in gene expression profiles among enterocytes. For example, immune genes are induced in enterocytes by microbiota and are negatively correlated with HNF4A-dependent genes, implying divergent transcriptional programs. We also find that IEC-specific deletion of Hnf4a leads to induction of

immune genes and enhanced protection from enterovirus infection, further confirming a negative relationship between HNF4A and immune programs. In accord, proteomic studies in gnotobiotic mouse IECs and pathway manipulations in human cells revealed significant interactions between HNF4A and immune signaling pathway

components. These data support our central hypothesis that HNF4A and immune pathways represent distinct microbiota-regulated enterocyte programs, and that HNF4A suppresses the immune program and thereby increases vulnerability to enterovirus infection. To test this, we will define the roles of HNF4A in mediating the

impacts of microbiota on IEC gene regulatory programs and enterovirus infection. We will also identify the molecular mechanisms by which HNF4A and immune pathways interact to regulate gene expression. The expected outcomes will vertically advance the field in several ways. First, they will provide an in-depth

understanding of how interactions between the microbiota and HNF4A determine IEC differentiation, gene expression, chromatin organization, and enterovirus infection. Second, they will yield unprecedented insights into the molecular mechanisms by which HNF4A and immune pathways communicate within the same cell.

These results are expected to have a positive impact because they would provide important fundamental knowledge about intestinal epithelial biology which could lead to the development of new diagnostic, prognostic, and therapeutic approaches for human diseases such as enteric virus infections.