Rotavirus impairs the biosynthesis of brush-border-associated dipeptidyl peptidase IV in human enterocyte-like Caco-2/TC7 cells

Abstract

SummaryRotavirus is the leading cause of severe dehydrating diarrhoea in infants and young children worldwide. This virus infects mature enterocytes in the small intestine, and induces structural and functional damage. In the present study, we have identified a new mechanism by which rotavirus impairs a brush border-associated intestinal protein. We show that infection of enterocyte-like Caco-2/TC7 cells by rhesus monkey rotavirus (RRV) impairs the biosynthesis of dipeptidyl peptidase IV (DPP IV), an important hydrolase in the digestion of dietary proline-rich proteins. We show that the enzyme activity of DPP IV was reduced, and that rearrangements of the protein occurred at the apical domain of the RRV-infected cells. Using pulse-chase experiments and cell surface immunoprecipitation, we have demonstrated that RRV infection did not affect the stability or apical targeting of DPP IV, but did induce a dramatic decrease in its biosynthesis. Using quantitative RT-PCR, we showed that RRV had no effect on the level of expression of DPP IV mRNA, suggesting that the observed decrease in the biosynthesis of the protein is related to an effect of the virus at the translational level.

Epithelial WNT Ligands Are Essential Drivers of Intestinal Stem Cell Activation

“…RV, a well-characterized, noninflammatory small intestinal viral pathogen, infects terminally differentiated mature cells, but not ISCs, in the small intestinal epithelium, resulting in watery diarrhea with vomiting, fever, and abdominal pain (Greenberg and Estes, 2009). RV infection alters the cytoskeleton and impairs host secretory pathways, leading to mislocalization of brush border enzymes and malabsorption, disruption of cell-cell junctions, increased permeability, cytotoxic effects, and activation of cell death (Ramig, 2004; Beau et al, 2007; Jourdan et al, 1998; Burns et al, 1995). Infection in adult mice is accompanied by a notable lack of villus blunting or epithelial ulceration, suggesting the induction of epithelial repair mechanisms that compensate and presumably replace the infected, lost cells (Ward et al, 1990; Burns et al, 1995; O’Neal et al, 1997; Blutt et al, 2012).…”

The Correlation Between the Expression of Differentiation Markers in Rat Small Intestinal Mucosa and the Transcript Levels of Schlafen 3

“…Colonic cell lines such as Caco-2 are frequently used to model small intestinal epithelial biology in vitro, 11,12 and “differentiation” becomes a moving target depending on the cell line, environment, stimuli, and end points under study. Although the normal colonic epithelium in vivo is certainly elaborately differentiated, normal colonocytes differ markedly from small bowel enterocytes in many ways, including the activity of glutamine synthetase 13 and the sensitivity of alkaline phosphatase 14 to mutagens 15 and its resistance to apoptosis.…”

Mn bioavailability by polarized Caco-2 cells: comparison between Mn gluconate and Mn oxyprolinate

“…The electric resistance evaluated the cell continuity; a transepithelial electric resistance ≥330 Ω/cm2 was required to obtain a tight monolayer [21]. In all the experiments the establishment of tight junctions [22] and the development of apical brush border [23] were determined by confocal microscopy in two filters randomly selected. These filters, upon fixation in 2% paraformaldehyde, were labeled with undiluted mouse-anti-human Junctional Adhesion Molecule (JAM, clone BV16, kind gift of Dr. Elisabetta Dejana, IFOM, The FIRC Institute of Molecular Oncology Foundation, Milan) [24] or mouse-anti-dipeptidyl-peptidase type IV (1:100, kind gift of Dr. Ralph Witzgall Institute of Anatomy and Cell Biology I, University of Heidelberg), both overnight at 4°C, revealed by goat-anti-mouse IgG AlexaFluor-594 (1:500, 45 min, Molecular Probes, Eugene, OR).…”