Neuroblastoma is the most common solid tumour in infants. It originates from committed sympathoadrenal progenitors of the embryonic neural crest (NC). MYCN, a proto-oncogene that normally regulates prenatal NC cell migration and expansion, is amplified in about 25% of primary tumours and is strongly associated with poor prognosis. Conditional human MYCN overexpression in the sympathoadrenal lineage in Th–MYCN-transgenic mice recapitulates neuroblastoma highly similar to the human disease. However, the molecular mechanism of MYCN-driven tumourigenesis is poorly understood. MYCN is known to regulate the expression of various microRNAs (miRNAs) in neuroblastoma, yet the exact roles of miRNAs in neuroblastoma tumourigenesis are unclear. The aim of this study is to identify novel miRNA-mRNA interactions that play roles in MYCN-driven neuroblastoma tumourigenesis. miRNA and mRNA expression profiling was performed on sympathetic ganglia from postnatal Th–MYCN+/+ mice during the pre-tumour stage and age-matched wild-type mice. Bayesian network with splitting-averaging analysis inferred a statistically significant miRNA-mRNA interaction network by integrating the sample matched miRNA/mRNA expression data. The putative targets of six miRNAs were examined further by enrichment analysis using GeneCodis3 and Molecular Signatures Database. One of these miRNAs is miR-204, which is significantly down-regulated in Th-MYCN+/+ ganglia compared to wild-type mice. The analysis of miRNA expression data from a 364-patients cohort revealed better overall and progression free survival with high miR-204 expression, and lower miR-204 expression in MYCN-amplified than in non-MYCN-amplified tumours. Importantly, miR-204 is up-regulated in human neuroblastoma cells upon MYCN siRNAs, suggesting negative regulation of miR-204 by MYCN. In addition, miR-204 down-regulates cell cycle enriched targets including E2F1, and MYCN. Consistently, miR-204 reduces neuroblastoma cell proliferation and colony forming capability. For the first time, our data suggest that miR-204 is a novel target of MYCN and a potential tumour suppressor in neuroblastoma tumourigenesis through down-regulating cell cycle-related genes.