RNA Isolation and qRT-PCR Analysis

RNA isolation and qRT-PCR analysis were performed as described previously^{8 (link),14 (link),50 (link)}. In short, RNA was isolated with a Machery-NucleoSpin RNA II kit (Bioké, Leiden, The Netherlands) according to the manufacturer’s instructions, and quantified using a Nanodrop ND-1000 (Wilmington, DE, USA). CDNA was prepared from total RNA using a cDNA Synthesis Kit (TAKARA BIO INC). Quantitative PCR was performed using Sensimix SYBRGreen (Applied Biosystems) or TaqMan (AXIN1; Hs00394718_m1, AXIN2; Hs00610344_m1, GAPDH; Hs02786624-g1) Gene Expression Assays (Applied Biosystems). Analyses were performed using the StepOne Real-Time PCR System and the StepOnev2.0 software (Applied Biosystems, Darmstadt, Germany). All expression levels are depicted relative to the expression of GAPDH. Primer sequences are provided in Supplemental Table S3.