Esophageal squamous cell carcinoma (ESCC) has one of the highest mortality rates worldwide. AU-rich element RNA-binding factor 1 (AUF1) is an established RNA-binding protein. AUF1 influences the process of development, apoptosis and tumorigenesis via interacting with adenylate-uridylate rich elements (AREs) bearing mRNAs. However, the clinical relevance of AUF1 and its biological function in ESCC progression have not been reported. In the present study, we first investigated the expression of AUF1 in the ESCC tissue samles and normal samples. We found a significantly higher expression of AUF1 in ESCC tissues than that in normal tissues and tumor adjacent tissues. The expression of AUF1 correlated with ESCC stage (P=0.011) and marginally correlated with lymph node metastasis (P=0.055) of ESCC patients. Silencing of AUF1 by an siRNA inhibited the proliferation and enhanced the apoptosis of ESCC cells. mRNA profiling by microarray analysis revealed that AUF1 knockdown affected 285 genes (fold change ≥2) that function in multiple pathways. GTP cyclohydrolase I (GCH1), the rate limiting enzyme for BH4 synthesis, was found to be downregulated. One of the AU-rich elements in the 3'UTR of GCH1 was found to be responsive to AUF1 expression by luciferase assay. Knockdown of GCH1 suppressed cell proliferation and colony formation of ESCC cells. The expression of AUF1 significantly correlated with that of GCH1 in ESCC tissues. Taken together, we demonstrated the overexpression of AUF1 in esophageal carcinoma and identified GCH1 as AUF1's effector for the proliferation of ESCC cells.