microRNA-34c is a novel target to treat dementias - PubMed (original) (raw)

. 2011 Sep 23;30(20):4299-308.

doi: 10.1038/emboj.2011.327.

Hope Y Agbemenyah, Roberto C Agis-Balboa, Roman M Stilling, Dieter Edbauer, Pooja Rao, Laurent Farinelli, Ivana Delalle, Andrea Schmitt, Peter Falkai, Sanaz Bahari-Javan, Susanne Burkhardt, Farahnaz Sananbenesi, Andre Fischer

Affiliations

microRNA-34c is a novel target to treat dementias

Athanasios Zovoilis et al. EMBO J. 2011.

Abstract

MicroRNAs are key regulators of transcriptome plasticity and have been implicated with the pathogenesis of brain diseases. Here, we employed massive parallel sequencing and provide, at an unprecedented depth, the complete and quantitative miRNAome of the mouse hippocampus, the prime target of neurodegenerative diseases such as Alzheimer's disease (AD). Using integrative genetics, we identify miR-34c as a negative constraint of memory consolidation and show that miR-34c levels are elevated in the hippocampus of AD patients and corresponding mouse models. In line with this, targeting miR-34 seed rescues learning ability in these mouse models. Our data suggest that miR-34c could be a marker for the onset of cognitive disturbances linked to AD and indicate that targeting miR-34c could be a suitable therapy.

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Conflict of interest statement

The authors declare the following conflict of interests. A patent application for the use of miR-34 to treat neurodegenerative diseases has been filed.

Figures

Figure 1

Figure 1

Massive parallel sequencing of small RNA libraries reveals the hippocampal miRNAome. (A) Contribution (in percent) of miRNAs to the total number of small non-coding RNAs in the range of 18–26 nt detected by sequencing. (B) Proportion of detected mature miRNAs, regarding the total number of known genes and miRNAs in miRBase. (C) Right panel: Distribution of miRNAs to different classes based to their sequence counts and contribution of each of these miRNA classes to the total miRNA sequence count. Left panel: Proportion of sequence counts per miRNA with respect to the total number of counts attributed to miRNAs in hippocampus. (D) Sequence counts of top ranking hippocampal miRNAs relative to the respective counts in whole brain −(log2 scale).

Figure 2

Figure 2

Genomic location of miR-34c and functional analysis of miR-34c targets. (A) Functional analysis of gene targets incorporating the miR-34c seed described in (B). A complete list of the genes that have been analysed is available in Supplementary Table SI (as defined in Targetscan Mouse, Release 5.1 12). Enrichment for Gene Ontology (GO) terms for biological processes and molecular functions at the fifth level as well as for the respective biological process terms from the PANTHER database was tested (top five terms for each GO term category are depicted). Analysis revealed a significant enrichment of miR-34c targets in terms strictly related to neuronal function, in contrast to other expressed or enriched in hippocampus miRNAs (see Supplementary Figure S3B–E). (B) UCSC browser (mm9) view of miR-34bc cluster. Genomic intervals of sequenced reads in mouse hippocampus are depicted in the first row in positions corresponding to miR-34c (left) and miR-34b (right) (third panel—UCSC miRNA track, as of January 2011). The next two panels depict conservation of the respective sequences among species (UCSC: placental mammal basewise conservation (up), multiz alignments of 30 vertebrates (down)). The last panel depicts the relative expression levels (according to sequence counts) of miR-34c and the other member of this cluster, miR-34b, which is minimally expressed. miR-34c presents high expression levels in mouse hippocampus and high evolutionary conservation, in line with an important role in cellular functions. (C) Percentage of each of miRNAs sharing the same with miR-34c seed and thus mRNA targets with regards to total amount of miRNAs detected in hippocampus targeting this seed. miR-34c represents >98% of miRNAs able to target this seed, revealing its almost exclusive control in regulation of mRNA targets of this group. miRNA seed as defined in Targetscan Mouse, Release 5.1 12. (D) miR-34c targets learning-associated genes. Enrichment of learning-associated genes upregulated 60 min after fear conditioning (see Supplementary Figure S4A) in seeds of miRNAs highly expressed in hippocampus.

Figure 3

Figure 3

High levels of miR-34c in mouse models for learning impairment. (A) Hippocampal miR-34c expression levels determined by qPCR in two mouse models for memory impairment, (left) 24-month-old mice (_n_=4; _P_=0.02), (right) 12-month-old APPPS1-21 mice (_n_=7, 6; _P_=0.01). (B) Left panel: 3- and 24-month-old C57Bl6J mice were subjected to contextual fear conditioning and freezing behaviour was analysed 24 h later during the memory test. Freezing was significantly reduced in 24-month-old mice (_n_=8/group, *P<0.05). Right panel: 12-month-old male APPPS1-21 mice were exposed to contextual fear conditioning. When compared with age-matched control littermates associative memory was impaired in APPPS1-21 mice. No difference in explorative behaviour or response to the foot-shock was observed among groups (data not shown). (C) SIRT1 protein levels are significantly reduced in the hippocampus of aged (_P_=0.02 versus young, _n_=4) and APPPS1-21 mice (_P_=0.01 versus wild type, _n_=7, 6). In line with a translational repression mode of action of miR-34c on SIRT1, Sirt1 mRNA levels were not affected. Error bars indicate s.e.m.

Figure 4

Figure 4

miR-34c is upregulated in AD patients. qPCR analysis was used to measure miR-34c levels in post-mortem tissue samples from AD patients and age-matched controls. (A) miR-34c levels (qPCR) are elevated in the hippocampus (*_P_=0.05) of human AD patients (_n_=6; 4 females, 2 males) when compared with age-matched controls (_n_=8, 2 females, 6 males). Five AD cases were diagnosed as Braack & Braack stage VI and one was diagnosed as Braack & Braack V. There was no significant difference for the age (B) or post-mortem delay (C) among groups. Error bars indicate s.e.m.

Figure 5

Figure 5

High levels of miR-34c are implicated with memory impairment. (A) Left panel: Experimental design for intrahippocampal administration of miR-34c mimic and the respective scramble miR. Right panel: Elevated hippocampal miR-34c levels after miR-34c mimic injection (_n_=5, 7; *_P_=0.05). (B) Upper panel, experimental design. Impaired learning in 3-month-old wild-type mice with high levels of miR-34c after treatment with the miRNA mimic (miR-34c) (_n_=6, *P<0.001). (C) Hippocampal SIRT1 levels correlate with miR-34c expression. miR-34c was injected into the hippocampus as shown in Figure 4A (_n_=4; *_P_=0.04). Sirt1 mRNA and protein levels were measured 24 h after fear conditioning. While elevation of miR-34c did not significantly affected Sirt1 mRNA levels (there was even a trend for increased Sirt1 mRNA, possible through compensatory mechanisms) we observed a dramatic effect in translation of this mRNA, leading to a significant repression of Sirt-1 protein levels. Error bars indicate s.e.m. (D) Relative SIRT1 protein/mRNA ratio for the experiment shown in (A).

Figure 6

Figure 6

miR-34c induces reduction of SIRT1 protein levels via its binding to SIRT1 3′UTR. (A) Experimental design. Upper panel: We designed miR-34c-Sirt1 protectors that hybridize to both miR-34c binding sites within the 3′ UTR of the Sirt1 mRNA. These protectors prevent miR34c mimic from binding to the Sirt1 mRNA. Lower panel: A control target protector oligonucleotide was used to exclude non-specific effects. (B) Upper panel: Experimental design. Lower panel: While intrahippocampal injection of miR-34c mimic along with the target protector negative control impaired associative learning in the contextual fear conditioning paradigm, co-injection with the miR-34c-Sirt1 protector rescues learning impairment (_n_=8, 8; *_P_=0.04). Mice co-injection with a scramble miR and the target protector negative control served as an additional control (_n_=8, 6; *_P_=0.04). (C) SIRT1 protein (left) and mRNA levels (right) in mice treated with the miR-34c mimic and miR-34c-Sirt1 protector compared with mice injected with miR-34c mimic along with the target protector negative control (*_P_=0.07; _n_=7, 5). Mmu, mus musculus. Error bars indicate s.e.m.

Figure 7

Figure 7

Targeting miR-34c seed rescues learning impairment in mouse model for AD. (A) Experimental design. (B) Impaired learning of 12-month-old APPPS1-21 mice (*_P_=0.001) is partially rescued after inhibition of the miR-34c activity (*_P_=0.04; _n_=7–8/group). (C) SIRT1 protein (left) and mRNA levels (left) in miR34 seed inhibitor-treated mice (*P<0.05; _n_=8). Error bars indicate s.e.m.

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References

    1. Alexiou P, Maragkakis M, Papadopoulos GL, Simmosis VA, Zhang L, Hatzigeorgiou AG (2010) The DIANA-mirExTra web server: from gene expression data to microRNA function. PLoS One 5: e9171. - PMC - PubMed
    1. Bak M, Silahtaroglu A, Møller M, Christensen M, Rath MF, Skryabin B, Tommerup N, Kauppinen S (2008) MicroRNA expression in the adult mouse central nervous system. RNA 14: 432–444 - PMC - PubMed
    1. Berchtold NC, Cribbs DH, Coleman PD, Rogers J, Head E, Kim R, Beach T, Miller C, Troncoso J, Trojanowski JQ, Zielke HR, Cotman CW (2008) Gene expression changes in the course of normal brain aging are sexually dimorphic. Proc Natl Acad Sci USA 105: 15605–15610 - PMC - PubMed
    1. Cannell IG, Bushell M (2010) Regulation of Myc by miR-34c: a mechanism to prevent genomic instability? Cell Cycle 9: 2726–2730 - PubMed
    1. Cao K, Chen-Plotkin AS, Plotkin JB, Wang LS (2010) Age-correlated gene expression in normal and neurodegenerative human brain tissues. PLoS One 5: e13098. - PMC - PubMed

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