The serine/threonine kinase UNC-51-like kinase 1 (ULK1) plays an essential role in autophagosome formation, although the exact molecular mechanism is unknown. The present study was first to investigate the clinical and prognostic significance of ULK1 in esophageal squamous cell carcinoma (ESCC). Protein and mRNA levels of ULK1 in normal esophageal epithelial cells, ESCC cell lines, paired ESCC lesions and the adjacent noncancerous tissues were examined using western blot and real-time RT-PCR. The results showed that only the ULK1 protein level was upregulated in ESCC samples compared with normal esophageal cells and tissues. Also, we found that protein stabilization of ULK1 was higher in ESCC cell lines. Furthermore, immunohistochemical staining of ULK1 was performed on the tissue microarray containing 248 ESCC and 51 normal esophageal tissues. A total of 70.2% ESCC specimens showed intensive expression of ULK1 in contrast to the undetectable expression of ULK1 in normal esophageal tissues. Statistical analysis revealed that ULK1 expression was significantly correlated with T status (P = 0.048). Moreover, patients with higher ULK1 expression were associated with shorter overall survival time. Multivariate analysis suggested that ULK1 expression and N status (P < 0.001) were independent prognostic indicators for the survival of patients. Functional studies showed that suppression of ULK1 expression in ESCC cell lines by specific small interfering RNA resulted in inhibition of cell proliferation and induction of apoptosis under starvation conditions. These findings provide evidence that ULK1 represents a novel and clinically useful biomarker for ESCC patients and plays an important role during the progression of ESCC. (Cancer Sci 2011; 102: 1568–1575)