Wnt3a stimulates Mepe, Matrix extracellular phosphoglycoprotein, expression directly by the activation of the canonical Wnt signaling pathway and indirectly through the stimulation of autocrine Bmp‐2 expression

Abstract

Matrix extracellular phosphoglycoprotein (MEPE) is a specific marker of mineralizing osteoblasts and osteocytes. Canonical BMP and Wnt signaling pathways are two of the strongest paracrine signals stimulating osteogenesis. Our previous results indicated that Mepe expression is stimulated by the BMP-2-signaling pathway. The specific aim of this study addressed whether Mepe expression is also controlled by Wnt signaling, and whether there is a cross-regulation between two major osteogenic signaling pathways. Treatment with Wnt3a, a canonical Wnt signaling stimulator, strongly enhanced Mepe mRNA expression. Knock-down of β-catenin with siRNA completely reversed Wnt3a-stimulated Mepe expression. The Mepe mRNA expression level was increased by overexpression of β-catenin and Lef-1, even in the absence of Wnt3a. Highly conserved Lef-1 response elements were identified in the mouse Mepe promoter. The direct binding of Lef-1 to these elements is critical for Mepe expression, indicating that Mepe is a direct target of canonical Wnt signaling. Meanwhile, we also found that Wnt3a treatment strongly stimulated Bmp-2 expression, and that the subsequent increase in Bmp-2 protein was determined in Wnt3a-treated conditioned medium (CM). Treatment of MC3T3-E1 cells with CM stimulated phosphorylation of the Smad1/5 proteins and their downstream Dlx5 mRNA expression. The CM-mediated increases of phospho-Smad and Dlx5 expression were not blocked completely by a Wnt3a antagonist, Dkk-1, but were almost completely suppressed by the addition of a Bmp-2 antagonist, Noggin. Collectively, Wnt3a stimulates Mepe transcription directly by a canonical Wnt signaling pathway through β-catenin and Lef-1 and indirectly through the activation of a Bmp-2 autocrine loop.