In this study, researchers performed extensive gene expression profiling experiments that identified three miRNA genes called miR-34s as candidate tumour suppressor genes
Scientists at Cold Spring Harbor Laboratory (CSHL), Howard Hughes Medical Institute (HHMI), and Applied Biosystems, have discovered that a family of small RNA molecules known as microRNAs (miRNAs) are components in a well-studied tumour suppressor network, the p53 pathway, that stops the growth of tumour cells in mice.
This tumour suppressor capability represents a newly discovered function for miRNAs.
The finding may also suggest new approaches for treating cancers.
The results of the team's two-year collaboration were published in the 28 June 2007 issue of the journal Nature.
Other co-authors were from Stony Brook University and Rosetta Inpharmatics.
MicroRNAs regulate gene expression, and have been implicated in a number of different cancers.
The researchers were able to detect significant changes in the amount of miR-34 miRNAs in mouse cells that had been constructed both with and without copies of the p53 gene, a gene that governs cell growth, suppressing tumour formation.
These findings helped them to explain the role of these miRNAs in tumour suppression.
Follow-up validation research with mice by scientists at CSHL revealed the cancer fighting function of the miRNAs.
"This study marks the first time that researchers have compared expression of microRNAs in cell samples that contain copies of the gene for p53 to expression of miRNA in cells in which the p53 gene copies have been removed," said Caifu Chen, a director of science at Applied Biosystems and a co-author of the study.
"Because of the success of this research, we now know that miRNAs have a tumour suppressor function".
Results of this study also showed that, in mouse tumour cells that contained a functioning p53 gene, expression levels of the three miR-34 genes increased compared to levels of the miRNAs in cells without the p53 gene.
The resulting miR-34 miRNAs then repressed the expression of other genes related to cell growth, which in cases of cancer, are involved in cell proliferation.
The researchers suggested that one way that p53 may suppress tumour growth is by inducing the expression of miR-34 genes, which then silence expression of various cell proliferation genes.
Consistent with their finding in mice tissue samples that miR-34 expression is regulated by p53, researchers found comparatively low levels of miR-34s in human tumours and cancer cell lines, which show a high frequency of p53 deficiency.
Scientists still do not fully understand what role miR-34 plays in the overall p53 tumour suppressor network.
However, they believe that there is great potential for altering miR-34 expression to harness the cancer fighting function of the p53 pathway and developing these miRNAs into therapeutic agents for treatment of cancer.
"Together, these data identify the miR-34 family of microRNAs as direct targets of p53 that play a key role in the p53 tumour suppressor network," said Lin He, Cold Spring Harbor Laboratory.
"Our finding reveals new details about how one of the most important and well-studied tumour suppressor networks prevents cancer formation".
The researchers in this study, who included Lin He, CSHL, and Caifu Chen, Applied Biosystems, used TaqMan MicroRNA Assays from Applied Biosystems for gene expression profiling experiments to identify differences between the amount of miR-34 genes expressed in samples of mouse cells that had both copies of the p53 gene and cells that had both copies removed.
TaqMan-based gene expression assays detect both the presence of target nucleic acids in a sample, and determine how much of that target is present in a sample.
Using TaqMan MicroRNA Assays, the researchers in this study were able to identify changes in amounts of miR-34 miRNAs in various cell samples, and determine the influence that the presence of p53 had on expression levels of the miRNAs.
Real-time PCR reactions, which are necessary to automate the assays, were performed using an Applied Biosystems 7900HT Real-Time PCR System.
Real-Time PCR systems automate TaqMan-based real-time PCR assays, making it possible for researchers to detect and quantify nucleic acid sequences.
The miR-34 genes were expressed at very low levels with, at most, only a few hundred copies of miRNA being present in cells.
The sensitivity and dynamic range of the TaqMan-based assays made it possible for researchers to detect and determine quantities of a broad range of miRNAs present in cell samples, including those miRNAs present at levels too low to detect and quantify using other methods.
Sensitivity is a measurement of the number of copies of target nucleic acid (DNA, mRNA, or miRNA) that must be present in a sample in order for the assay to positively identify the target.
"The sensitivity and dynamic range of the TaqMan MicroRNA Assays made it possible for us to precisely measure the levels of miR-34 genes expressed in cells with and without copies of the p53 gene," said Dr He.
