Cellular polarity asymmetrically functionalizes pathogen recognition receptor-mediated intrinsic immune response in human intestinal epithelium cells

Abstract

50Intestinal epithelial cells (IECs) act as a physical barrier separating the 51 commensal-containing intestinal tract from the sterile interior. These cells have 52 found a complex balance allowing them to be prepared for pathogen attacks 53 while still tolerating the presence of bacteria/viral stimuli present in the lumen 54 of the gut. Using primary human IECs, we probed the mechanisms, which allow 55 for such a tolerance. We discovered that viral infection emanating from the 56 basolateral side of IECs elicited a stronger intrinsic immune response as 57 compared to lumenal apical infection. We determined that this asymmetric 58 immune response was driven by the clathrin-sorting adapter AP-1B which 59 mediates the polarized sorting of Toll-like receptor 3 (TLR3) toward the 60 basolateral side of IECs. Mice and human IECs lacking AP-1B showed an 61 exacerbated immune response following apical stimulation. Together these 62 results suggest a model where the cellular polarity program plays an integral 63 role in the ability of IECs to partially tolerate apical commensals while remaining 64 fully responsive against invasive basolateral pathogens. 65 66 Keywords: Intrinsic innate immunity, type III interferon, TLR3, cellular polarity, 67 clathrin, AP-1B, lambda interferon, enteric viruses, organoids, intestinal 68 epithelial cells 69 70 71

Critical Role of Type III Interferon in Controlling SARS-CoV-2 Infection in Human Intestinal Epithelial Cells

“…We found that $10% of the cells are infected in a human-colon-derived organoid, leading to a modest but significant increase in viral genome copy number (Figure 4F) and de novo infectious virus particles (Figure 4J) over time. This modest replication of SARS-CoV-2 is explained by the fact that (1) only a small fraction of the cells are infected by SARS-CoV-2, and (2) as shown in this work, IFNs are potent inhibitors of SARS-CoV-2 replication (Figures 2 and 3), and organoids are highly immunoresponsive upon viral infection (Stanifer et al, 2020) (Figure 4G). Our findings are in agreement with recent work from Lamers et al showing that infection of human intestinal organoids by SARS-CoV-2 induces a strong immune response characterized by a strong expression of ISGs (Lamers et al, 2020).…”

“…This highlights that host-pathogen interaction should be considered in a tissue-specific manner, as different cellular responses and viral countermeasures might be established among the lung, gut, and other organs. Interestingly, in human colon organoids, we observed that only type III IFN is made upon SARS-CoV-2 infection, although human intestinal organoids are capable of making both type I and III IFN upon enteric virus infection (Pervolaraki et al, 2018;Stanifer et al, 2020). The lack of type I induction could be due to a kinetic delay of type I IFN production compared to type III IFN but this will need to be carefully addressed in further experiments.…”

“…Intact ultrastructural organization and differentiation to all cell types (e.g., enterocytes, Goblet cells, enteroendocrine cells, and stem cells) was confirmed using confocal fluorescent microscopy (Fig- ure 4A) and qRT-PCR against cell-type-specific transcripts (Figure 4B). As quantification of the number of infected cells in 3D organoids is very challenging, we exploited previously established protocols to differentiate and infect human intestinal organoids in two dimensions with viruses (Ettayebi et al, 2016;Stanifer et al, 2020). Nondifferentiated organoids were seeded on human-collagen-coated iBIDI chambers.…”

Using Diverse Model Systems to Define Intestinal Epithelial Defenses to Enteric Viral Infections

“…These differences likely affect the responsiveness of the epithelium to the microbiota and result in differences in microbiota-mediated tonic signaling between the mouse and human intestine (Hamonic et al, 2018;Price et al, 2018). Indeed, studies in mice have shown that TLR stimulation of enterocytes apically versus basolaterally results in different responses (Lee et al, 2006;Stanifer et al, 2019). Moreover, systemic flagellin, but not LPS, protected mice against enteric rotavirus in a TLR5-and NLR family CARD domain-containing 4 (NLRC4)dependent manner (Uchiyama et al, 2014).…”

“…Moreover, systemic flagellin, but not LPS, protected mice against enteric rotavirus in a TLR5-and NLR family CARD domain-containing 4 (NLRC4)dependent manner (Uchiyama et al, 2014). This is likely due to the different immune responses as is seen for apical versus basolateral stimulation of TLRs on intestinal epithelial cells (IECs) (Lee et al, 2006;Stanifer et al, 2019). For example, TLR5 is located on the basolateral side of IECs, which would make it only responsive to systemic flagellin (Rhee et al, 2005).…”