Fibroblast Growth Factor Receptor-3 (FGFR-3) Regulates Expression of Paneth Cell Lineage-specific Genes in Intestinal Epithelial Cells through both TCF4/β-Catenin-dependent and -independent Signaling Pathways

Leigh Bradley,Alda Vidrich,Steven M Cohn,Brooks Brodrick,Jenny M Buzan,Edith Porter

doi:10.1074/jbc.m111.229252

Abstract

Fibroblast growth factor receptor-3 (FGFR-3) expression in the developing intestine is restricted to the undifferentiated epithelial cells within the lower portion of the crypt. We previously showed that mice lacking functional FGFR-3 have a significant decrease in the number of Paneth cells in the small intestine. Here, we used Caco2 cells to investigate whether FGFR-3 signaling can directly modulate expression of Paneth cell differentiation markers through its effects on TCF4/β-catenin or through other signaling pathways downstream of this receptor. Caco2 cells treated with FGFR-3 ligands or expressing FGFR-3(K650E), a constitutively active mutant, resulted in a significantly increased expression of genes characteristic of mature Paneth cells, including human α-defensins 5 and 6 (HD5 and HD6) and Paneth cell lysozyme, whereas enterocytic differentiation markers were reduced. Activation of FGFR-3 signaling sustained high levels of β-catenin mRNA expression, leading to increased TCF4/β-catenin-regulated transcriptional activity in Caco2 cells. Sustained activity of the TCF4/β-catenin pathway was required for the induction of Paneth cell markers. Activation of the MAPK pathway by FGFR-3 is also required for the induction of Paneth cell markers in addition to and independent of the effect of FGFR-3 on TCF4/β-catenin activity. These studies suggest that coordinate activation of multiple independent signaling pathways downstream of FGFR-3 is involved in regulation of Paneth cell differentiation.

Highlights

Lysozyme, phospholipase A2, ␣-defensins in humans and cryptdins in mice, and RegIII␥, that are secreted into the lumen and neutralize invading microorganisms [3, 6] Consistent with their role in innate immunity in the intestine, several studies have shown that dysregulation of Paneth cell function is associated with chronic intestinal inflammation in animal models of ileitis and with Crohn’s disease in humans [7, 8]
We report that activation of Fibroblast growth factor receptor-3 (FGFR-3) in Caco2 cells by ligands or by expression of a ligand-independent, constitutively active mutant of FGFR-3, FGFR-3K650E, results in the coordinately induced expression of multiple genes characteristic of mature intestinal Paneth cells
Our current results suggest that the effects of FGFR-3 on Paneth cell differentiation require independent activation of multiple signaling pathways downstream of FGFR-3 in undifferentiated epithelial progenitors within the crypt epithelium

Summary

Introduction

Lysozyme, phospholipase A2, ␣-defensins in humans and cryptdins in mice, and RegIII␥, that are secreted into the lumen and neutralize invading microorganisms [3, 6] Consistent with their role in innate immunity in the intestine, several studies have shown that dysregulation of Paneth cell function is associated with chronic intestinal inflammation in animal models of ileitis and with Crohn’s disease in humans [7, 8]. By postnatal day 21, FGFR-3Ϫ/Ϫ mice show a marked deficit in a number of Paneth cell markers and a significant reduction in the number of lysozyme-positive Paneth cells These mice have a reduced number of intestinal crypts compared with wild-type mice that is a reflection of diminished numbers of crypt stem cells [13]. These data suggest that FGFR-3-mediated signaling events play a role in the developmental regulation of the stem cell compartment as well as in the commitment of progenitor cells to Paneth cell lineage-specific differentiation. Wnt is the most well known regulator of TCF4/␤-catenin signaling, other growth factors, such as the FGFR-3 ligands FGF2 and FGF9, can mediate signaling via this pathway [13, 20]

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Conclusion