DS is a complex syndrome of genetic origin with multiple and variable clinical features [3
]. These phenotypic outcomes seem to be the product of dosage effects of multiple genes affecting developmental processes and functions and not a simple one gene, one phenotype explanation [7
]. To date, the contribution of miRNAs in DS has not been investigated. For the first time we demonstrate that mature miR99a, let-7c, miR-125b-2, miR-155 and miR- 802, which are encoded by genes harbored on Hsa21, are over-expressed in human fetal hippocampus and heart samples from individuals with DS. As previously described, the end result of miRNA-mediated gene regulation is a reduction in the total amount of target protein that is produced [8
], we speculate that Trisomy 21-induced over-expression of Hsa21-derived miRNAs, results in the decreased expression of specific target proteins and contributes, in part, to features of the DS phenotype.
It is estimated that 30% to 90% of human genes are regulated by miRNAs [22
]. Recently, several general principles regarding miRNAs and target-gene regulation have emerged. First, each miRNA can potentially regulate a large number of protein-coding genes. Second, many miRNAs potentially act in combination to regulate the same target genes. This allows for combinatorial regulation, where synergy between individual target sites has been demonstrated [9
]. Finally, predicted miRNA target genes are not restricted to a particular functional category or biological pathway, but rather are involved in a wide variety of biological processes. Therefore, based on these parameters the over-expression of the five Hsa21-derived miRNAs in DS individuals may result in the aberrant expression of a myriad of critical proteins in a variety of tissues.
MiRNA-mediated regulation of gene expression results when a miRNA binds to its recognition element within the 3′-UTR of a target mRNA and suppresses its translation or initiates its degradation (8
). The algorithms used to predict miRNA/mRNA targets typically develop scoring schemes based on sequence complementarity, free energy calculations of RNA:duplex formation, and phylogenetic conservation. For example, if the bioinformatic algorithm “TargetScan” [23
] is utilized (http://www.targetscan.org
) to predict Hsa21-derived miRNA/mRNA target sites, the following information is obtained: miR-99a, 36 putative mRNA targets; let-7c, 616 putative mRNA targets; miR-125b-2, 488 putative mRNA targets; miR-155, 240 putative mRNA targets, and miR-802, 190 putative mRNA targets. Current algorithms utilize conservation of miRNA/mRNA target sites across species as an important parameter; importantly however, the conservation of a miRNA target site is not a requirement for a functional miRNA. Therefore, the actual number of Hsa21-derived miRNA/mRNA targets is much greater than the number of in silico
predicted recognition elements denoted above. Although Hsa21-derived miRNAs may regulate large numbers of mRNA targets, therapeutically this is not a disadvantage since inhibition or knock-down of these over-expressed miRNAs should normalize the expression level of all miRNA/mRNA targets back to non-trisomic 21 levels. Importantly, recent studies utilizing locked nucleic acid (LNA)-modified anti-sense oligonucleotides (antimiRs), which are complementary to specific miRNAs, have demonstrated that LNA antagonists have high metabolic stability, specificity, efficacy, and potency [24
] and are therefore well suited for the development of novel therapeutics for individuals with DS.