A number of studies have shown that increased APP levels, resulting from either a genomic locus duplication or alteration in APP regulatory sequences, can lead to development of early-onset dementias, including Alzheimer's disease (AD). Therefore, understanding how APP levels are regulated could provide valuable insight into the genetic basis of AD and illuminate novel therapeutic avenues for AD. Here we test the hypothesis that APP protein levels can be regulated by miRNAs, evolutionarily conserved small noncoding RNA molecules that play an important role in regulating gene expression. Utilizing human cell lines, we demonstrate that miRNAs hsa-mir106a and hsa-mir-520c bind to their predicted target sequences in the APP 3'UTR and negatively regulate reporter gene expression. Over-expression of these miRNAs, but not control miRNAs, results in translational repression of APP mRNA and significantly reduces APP protein levels. These results are the first to demonstrate that levels of human APP can be regulated by miRNAs. ResultsAccumulating evidence suggests that increased expression of the amyloid precursor protein gene (APP) increases Alzheimer's disease (AD) risk. The resulting increase in APP protein levels results in increased A levels, leading to synaptic dysfunction, neurodegeneration and, eventually, cognitive decline.APP levels can be regulated at the genomic, transcriptional or translational level. At the genomic level, Down's Syndrome (Trisomy 21) patients have three copies of the APP gene and develop AD symptoms early in life [1]. Similarly, duplication of the APP locus, in the absence of a full trisomy 21, also leads to early-onset AD [2]. Dysregulation of APP transcription can also increase the risk of AD. Genetic variants in the APP promoter increase APP transcription by ~2-3 fold and have been reported to increase AD risk [3]. Growth factors have been reported to control APP mRNA half-life [4]. These growth factors effects are dependent on a 29 bp sequence in the APP 3' UTR [4,5]. APP translation is also regulated; for example, IL-1 can induce an increase in APP translation [6]. IL-1 is a proinflammatory cytokine and genetic variants have been linked to increased AD risk [7,8]. Taken together, these findings provide strong evidence that increased APP levels increase AD risk.

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BackgroundMicroRNAs (miRNAs: a class of short non-coding RNAs) are emerging as important agents of post transcriptional gene regulation and integral components of gene networks. MiRNAs have been strongly linked to stem cells, which have a remarkable dual role in development. They can either continuously replenish themselves (self-renewal), or differentiate into cells that execute a limited number of specific actions (pluripotence).Methodology/Principal FindingsIn order to identify novel miRNAs from narrow windows of development we carried out an in silico search for micro-conserved elements (MCE) in adult tissue progenitor transcript sequences. A plethora of previously unknown miRNA candidates were revealed including 545 small RNAs that are enriched in embryonic stem (ES) cells over adult cells. Approximately 20% of these novel candidates are down-regulated in ES (Dicer −/−) ES cells that are impaired in miRNA maturation. The ES-enriched miRNA candidates exhibit distinct and opposite expression trends from mmu-mirs (an abundant class in adult tissues) during retinoic acid (RA)-induced ES cell differentiation. Significant perturbation of trends is found in both miRNAs and novel candidates in ES (GCNF −/−) cells, which display loss of repression of pluripotence genes upon differentiation.Conclusion/SignificanceCombining expression profile information with miRNA target prediction, we identified miRNA-mRNA pairs that correlate with ES cell pluripotence and differentiation. Perturbation of these pairs in the ES (GCNF −/−) mutant suggests a role for miRNAs in the core regulatory networks underlying ES cell self-renewal, pluripotence and differentiation.

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