Previous studies have indicated the important roles of MYCN in tumorigenesis and progression of neuroblastoma (NB), the most common extracranial solid tumor derived from neural crest in childhood. However, the regulatory mechanisms of MYCN expression in NB still remain largely unknown. In this study, through mining public microarray databases and analyzing the cis-regulatory elements and chromatin immunoprecipitation data sets, we identified CCCTC-binding factor (CTCF) as a crucial transcription factor facilitating the MYCN expression in NB. RNA immunoprecipitation, RNA electrophoretic mobility shift assay, RNA pull down and in vitro binding assay indicated the physical interaction between CTCF and MYCN opposite strand (MYCNOS), a natural noncoding RNA surrounding the MYNC promoter. Gain- and loss-of-function studies revealed that MYCNOS facilitated the recruitment of CTCF to its binding sites within the MYCN promoter to induce chromatin remodeling, resulting in enhanced MYCN levels and altered downstream gene expression, in cultured NB cell lines. CTCF cooperated with MYCNOS to suppress the differentiation and promote the growth, invasion and metastasis of NB cells in vitro and in vivo. In clinical NB tissues and cell lines, CTCF and MYCNOS were upregulated and positively correlated with MYCN expression. CTCF was an independent prognostic factor for unfavorable outcome of NB, and patients with high MYCNOS expression had lower survival probability. Taken together, these results demonstrate that CTCF cooperates with noncoding RNA MYCNOS to exhibit oncogenic activity that affects the aggressiveness and progression of NB through transcriptional upregulation of MYCN.

SummaryA suite of newly discovered sucrose transporter genes, PsSUF1, PsSUF4, PvSUT1 and PvSUF1, were isolated from the coats of developing pea (Pisum sativum L.) and bean (Phaseolus vulgaris L.) seeds. Sequence analysis indicated that deduced proteins encoded by PsSUF1, PvSUT1 and PvSUF1 clustered in a separate sub-group under sucrose transporter Clade I, whereas the deduced protein encoded by PsSUF4 clustered in Clade II. When expressed in yeast, these genes were shown to encode sucrose transporters with apparent Michaelis Menten constant (K m ) values ranging from 8.9 to 99.8 mM. PvSUT1 exhibited functional characteristics of a sucrose/H þ symporter. In contrast, PsSUF1, PvSUF1 and PsSUF4 supported the pH-and energy independent transport of sucrose that was shown to be bi-directional. These transport properties, together with that of counter transport, indicated that PsSUF1, PvSUF1 and PsSUF4 function as carriers that support the facilitated diffusion of sucrose. Carrier function was unaffected by diethylpyrocarbonate and by maltose competition, suggesting that the sucrose binding sites of these transporters differed from those of known sucrose/H þ symporters. All sucrose transporters were expressed throughout the plant and, of greatest interest, were coexpressed in cells considered responsible for sucrose efflux from seed coats. The possible roles played by the novel facilitators in sucrose efflux from seed coats are discussed.

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