MicroRNAs (miRNAs), noncoding RNAs 21-25 nucleotides in length, regulate gene expression primarily at the posttranscriptional level. Growing evidence suggests that miRNAs are aberrantly expressed in many human cancers, and that they play significant roles in carcinogenesis and cancer progression. A search for miRNAs with a tumor-suppressive function in esophageal squamous cell carcinoma (ESCC) was performed using the miRNA expression signatures obtained from ESCC clinical specimens. A subset of 15 miRNAs was significantly downregulated in ESCC. A comparison of miRNA signatures from ESCC and our previous report identified 4 miRNAs that are downregulated in common (miR-145, miR-30a-3p, miR-133a and miR-133b), suggesting that these miRNAs are candidate tumor suppressors. Gain-of-function analysis revealed that 3 transfectants (miR-145, miR-133a and miR-133b) inhibit cell proliferation and cell invasion in ESCC cells. These miRNAs (miR-145, miR-133a and miR-133b), which have conserved sequences in the 3 0 UTR of FSCN1 (actin-binding protein, Fascin homolog 1), inhibited FSCN1 expression. The signal from a luciferase reporter assay was significantly decreased at 2 miR-145 target sites and 1 miR-133a/b site, suggesting both miRNAs directly regulate FSCN1. An FSCN1 loss-of-function assay found significant cell growth and invasion inhibition, implying an FSCN1 is associated with ESCC carcinogenesis. The identification of tumor-suppressive miRNAs, miR-145, miR-133a and miR-133b, directly control oncogenic FSCN1 gene. These signal pathways of ESCC could provide new insights into potential mechanisms of ESCC carcinogenesis.Human esophageal cancer occurs worldwide with a variable geographic distribution. The disease ranks eighth in order of occurrence and sixth as the leading cause of cancer mortality, affecting men more than women. 1 There are 2 main forms, each with distinct etiologic and pathologic characteristics: esophageal squamous cell carcinoma (ESCC) and adenocarcinoma. ESCC is the most frequent subtype of esophageal cancer, although the incidence of adenocarcinoma in the western world is increasing faster than other malignancies. ESCC is one of the most aggressive and lethal malignancies in eastern Asia. Because most cases of ESCC are not diagnosed until the disease is at an advanced stage, the overall 5-year survival rate is very poor.2-4 Recently, the combination of chemotherapy and radiotherapy, alone or as an adjunct to surgery, has improved the prognosis of ESCC patients. 5,6 Research over the last 20 years has identified a number of oncogenic and tumor-suppressor proteins that are associated with the induction of ESCC. 7,8 However, molecular indicators of the origin of cellular deregulation in ESCC have not been identified. Elucidation of the molecular pathways involved in ESCC carcinogenesis could lead to improvements in disease diagnosis and therapy.MicroRNAs (miRNAs) are a class of naturally occurring small (21-25 nucleotides) noncoding RNAs. Mature miRNAs play important regulatory roles in cell growth, p...