Institute of Information Science Academia Sinica

MicroRNAs (miRNAs) are a family of small non-coding RNA molecules 
of about 20-23 nucleotides in length, which negatively regulate 
protein-coding genes at posttranscriptional manner. Computational 
analyses have been estimated that the number of microRNA in human 
genome may over 1000 genes which may regulate as many as 30% of human
protein coding genes. The expression levels of microRNA vary 
greatly among tissues and under different physiological and 
pathological conditions. Previous studies have shown that some 
microRNAs function as tumor suppressors while other microRNAs 
behave as oncogenes.

We describe a simple, robust, inexpensive method for microRNA 
quantification using stem-loop RT followed by SYBR green qPCR 
analysis. The assay is sensitive and has a linear range for microRNA 
concentrations that span more than 8 orders of magnitude. We used 
the method to quantify the expression levels of 270 human miRNAs
in 8 pairs of nasopharyngeal carcinoma (NPC) and adjacent 
non-tumor tissue (NT) and identified several miRNAs whose 
expression levels were significantly altered in NPC samples. Among
the differentially expressed miRNA is hsa-miR-9, which showed a 
more than 5-fold reduction in NPC samples compared to normal tissues.
Sequence analysis revealed the presence of CpG islands in the
promoter region of hsa-miR-9-1. Bisulphite-sequencing data 
confirmed that the promoter region of hsa-miR-9-1 was 
hypermethylated in NPC cell lines as well as in primary tissues from 
patient with NPC. In NPC cell lines, the expression levels of 
has-miR-9 were significantly increased after treatment with the
demethylating agent, 5-aza-2’-deoxycytidine. These results 
demonstrate that the promoter region of hsa-miR-9-1 is 
hypermethylated in NPC tissues and suggest promoter methylation may 
be responsible for the down-regulation of hsa-miR-9 in NPC. Study 
is currently on-going to investigate the biological function and
pathological significance of hsa-miR-9 in NPC.