Small Intestine Inflammation in Roquin-Mutant and Roquin-Deficient Mice

Abstract

Roquin, an E3 ubiquitin ligase that localizes to cytosolic RNA granules, is involved in regulating mRNA stability and translation. Mice that have a M199R mutation in the Roquin protein (referred to as sanroque or Roquinsan/san mice) develop autoimmune pathologies, although the extent to which these occur in the intestinal mucosa has not been determined. Here, we demonstrate that Roquinsan/san mice reproducibly develop intestinal inflammation in the small intestine but not the colon. Similarly, mice generated in our laboratory in which the Roquin gene was disrupted by insertion of a gene trap cassette (Roquingt/gt mice) had small intestinal inflammation that mimicked that of Roquinsan/san mice. MLN cells in Roquinsan/san mice consisted of activated proliferating T cells, and had increased numbers of CD44hi CD62Llo KLRG1+ short-lived effector cells. Proportionally more small intestinal intraepithelial lymphocytes in Roquinsan/san mice expressed the ICOS T cell activation marker. Of particular interest, small intestinal lamina propria lymphocytes in Roquinsan/san mice consisted of a high proportion of Gr-1+ T cells that included IL-17A+ cells and CD8+ IFN-γ+ cells. Extensive cytokine dysregulation resulting in both over-expression and under-expression of chemotactic cytokines occurred in the ileum of Roquinsan/san mice, the region most prone to the development of inflammation. These findings demonstrate that chronic inflammation ensues in the intestine following Roquin alteration either as a consequence of protein mutation or gene disruption, and they have implications for understanding how small intestinal inflammation is perpetuated in Crohn's disease (CD). Due to the paucity of animal models of CD-like pathophysiology in the small intestine, and because the primary gene/protein defects of the Roquin animal systems used here are well-defined, it will be possible to further elucidate the underlying genetic and molecular mechanisms that drive the disease process.

Hematopoietic not systemic impairment of Roquin expression accounts for intestinal inflammation in Roquin-deficient mice

“…Because Rc3h1 gt/gt mice made in our laboratory were generated by a random insertion of a gene-trap into the murine genome, the likelihood of simultaneous insertion into both the Rc3h1 and Rc3h2 genes was extremely remote. We therefore conclude, based on the studies here and our previous work7, that ablation of only the Rc3h1 gene is by itself sufficient to render an autoimmune phenotype in mice. It should be noted, however, that Roquin-1 and Roquin-2 expression were reported to be variable in mice throughout various immunological compartments21.…”

“…Collectively, these findings demonstrate that mice generated from BM of animals with targeted disruption of the Rc3h1 gene develop extensive inflammation similar to that of Rc3h1 san/san and Rc3h1 gt/gt mice7. Because, the Rc3h1 gene was unaltered in non-hematopoietic cells of Rc3h1 gt/gt → NL chimeras, the pathology which ensued in Rc3h1 gt/gt mice was attributable to the inflammatory response generated from cells of the immune system.…”

“…Sanroque mice ( Rc3h1 san/san ), which have an M199R mutation in the Roquin protein, develop extensive chronic inflammation consisting of lymphadenopathy, splenomegaly, thrombocytopenia, necrotizing hepatitis, increased CD4 + follicular T cells and expression of the inducible costimulator (ICOS) marker of T cell activation23456. Additionally, we recently demonstrated that Rc3h1 san/san mice have increased ICOS and OX40 expression in mesenteric lymph node T cells, and that a Crohn's disease-like chronic small intestinal inflammatory response develops in those animals7. The latter is of particular interest given the paucity of animal models of small intestine inflammation.…”

“…Because mice with a disrupted Rc3h1 gene ( Rc3h1 gt/gt mice) have physiological changes that extend beyond the immune system, the intestinal inflammatory response and tissue destruction observed in those animals7 could be caused by non-immunological perturbations, e.g., Roquin-mediated changes in epithelial cell permeability or effects directed at other non-hematopoietic tissues. To address this, we have used bone marrow (BM) radiation chimeras generated by injecting Rc3h1 gt/gt BM into irradiated normal (NL) mice ( Rc3h1 gt/gt → NL chimeras) to determine whether the intestinal inflammation in Rc3h1 gt/gt mice was due to a disruption of the Rc3h1 gene in cells of the immune system (in which case inflammation would be evident), or whether it required systemic Roquin impairment (in which case inflammation would be absent).…”

Inducible T-cell co-stimulator ligand (ICOSL) blockade leads to selective inhibition of anti-KLH IgG responses in subjects with systemic lupus erythematosus

“…14 16 18-23 Dysregulation of the pathway through a point mutation in RC3H1 (roquin 1), a repressor of ICOS, leads to the overexpression of ICOS and the development of a lupus-like systemic autoimmune disease in mice. [24][25][26] Pharmacological blockade and use of knockout rodents in animal models of autoimmune diseases have demonstrated the potential for therapeutic intervention of this pathway in human autoimmune diseases such as systemic lupus erythematosus (SLE), asthma and rheumatoid arthritis (RA). [27][28][29][30] SLE is a multisystem autoimmune disease of unknown cause with diverse clinical manifestations that disproportionately affects minorities and women of childbearing potential.…”