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Background: Increased protein N-glycosylation and aberrant Wnt signaling are features of oral cancer. Results: Overexpression of pro-migratory protein CTHRC1 is due to hyperglycosylation and transcriptional activation by canonical Wnt. Conclusion: N-Glycosylation collaborates with canonical Wnt to induce CTHRC1 and drive OSCC cell migration. Significance: Elucidating how N-glycosylation impacts tumor-promoting proteins is critical to understand cancer development and progression.

The transcription start site of the collagen ␣2(1) gene (COL1A2) has a sequence-specific binding site for a DNA methylation-responsive binding protein called regulatory factor for X-box 1 (RFX1) (Sengupta, P. K., Erhlich, M., and Smith, B. D. (1999) J. Biol. Chem. 274, 36649 -36655). In this report, we demonstrate that RFX1 forms homodimers as well as heterodimers with RFX2 spanning the collagen transcription start site. Methylation at ؉7 on the coding strand increases RFX1 complex formation in gel shift assays. Methylation on the template strand, however, does not increase RFX1 complex formation. DNA from human fibroblasts contains minimal methylation on the coding strand (<4%) with variable methylation on the template strand. RFX1 acts as a repressor of collagen transcription as judged by in vitro transcription and co-transfection assays with an unmethylated collagen promoter-reporter construct. In addition, an RFX5 complex present in human fibroblasts interacts with the collagen RFX site, which is not sensitive to methylation. This is the first demonstration of RFX5 complex formation on a gene other than major histocompatibility complex (MHC) promoters. Also, RFX5 represses transcription of a collagen promoterreporter construct in rat fibroblasts that have no detectable RFX5 complex formation or protein.