MicroRNA-138 is a potential regulator of memory performance in humans

Abstract

Genetic factors underlie a substantial proportion of individual differences in cognitive functions in humans, including processes related to episodic and working memory. While genetic association studies have proposed several candidate "memory genes," these currently explain only a minor fraction of the phenotypic variance. Here, we performed genome-wide screening on 13 episodic and working memory phenotypes in 1318 participants of the Berlin Aging Study II aged 60 years or older. The analyses highlight a number of novel single nucleotide polymorphisms (SNPs) associated with memory performance, including one located in a putative regulatory region of microRNA (miRNA) hsa-mir-138-5p (rs9882688, P-value = 7.8 × 10(-9)). Expression quantitative trait locus analyses on next-generation RNA-sequencing data revealed that rs9882688 genotypes show a significant correlation with the expression levels of this miRNA in 309 human lymphoblastoid cell lines (P-value = 5 × 10(-4)). In silico modeling of other top-ranking GWAS signals identified an additional memory-associated SNP in the 3' untranslated region (3' UTR) of DCP1B, a gene encoding a core component of the mRNA decapping complex in humans, predicted to interfere with hsa-mir-138-5p binding. This prediction was confirmed in vitro by luciferase assays showing differential binding of hsa-mir-138-5p to 3' UTR reporter constructs in two human cell lines (HEK293: P-value = 0.0470; SH-SY5Y: P-value = 0.0866). Finally, expression profiling of hsa-mir-138-5p and DCP1B mRNA in human post-mortem brain tissue revealed that both molecules are expressed simultaneously in frontal cortex and hippocampus, suggesting that the proposed interaction between hsa-mir-138-5p and DCP1B may also take place in vivo. In summary, by combining unbiased genome-wide screening with extensive in silico modeling, in vitro functional assays, and gene expression profiling, our study identified miRNA-138 as a potential molecular regulator of human memory function.

Keywords: DCP1B; GWAS; episodic memory; genome-wide association study; hsa-mir-138-5p; microRNA; working memory.

Figure 1

Predicted miRNA binding sites in the DCP1B transcripts. (A) Rs112215626 is located in the seed region of hsa-miR-4775. The alternative (G) allele of rs112215626 (highlighted in red) may alter binding affinity of hsa-miR-4775 to the DCP1B 3′UTR and therefore alter protein levels. (B) Rs1044950 is located in the seed region of hsa-miR-138-5p. The alternative T allele of the rs1044950 (highlighted in red) may alter binding affinity of hsa-miR-138-5p to the DCP1B 3′UTR and subsequently alter protein levels.

Figure 2

In vitro effects of rs1044950 and rs112215262 on miRNA-to-mRNA binding and gene expression. The bar charts show the normalized Renilla luciferase expression in constructs containing DCP1B 3′UTR and corresponding SNP alleles. Depicted are the mean Renilla luciferase intensities and the standard errors relative to the control luciferase intensities of the construct co-transfected with the non-targeting miRNA control (corresponding to the horizontal line): (A) for transcript ENST00000541700 containing the reference (G) or alternative (A) allele of rs1044950 and co-transfected with has-miR-138-5p into HEK293 and SH-SY5Y cells. The relative mean luciferase luminescence of the construct containing the G and the A allele was 0.585 (±0.0335) and 0.703 (±0.0417) in HEK293 cells, and 0.880 (±0.0256) and 0.985 (±0.0513) in SH-SY5Y cells. (B) for transcript ENST00000540622 containing the reference (T) or alternative (C) allele of rs112215626 and co-transfected with hsa-miR-4775 into HEK293 and SH-SY5Y cells. The relative mean luciferase luminescence of the construct containing T and the C allele was 0.903 (±0.0692) and 0.931 (±0.0382) in HEK293 cells, and 1.056 (±0.0582) and 1.041 (±0.0185) in SH-SY5Y cells.

Figure 3

Expression profile of hsa-miR-138-5p and DCP1B in autopsy brain tissues of three deceased human probands. (A) Amplification plot of qPCR experiments in three frontal cortices (highlighted in purple) and three hippocampi (highlighted in green). Hsa-miR-138-5p was expressed in both frontal cortex (FC) and hippocampus (HC; C_T ~ 19). One of two NTCs (non-template control, highlighted in red) showed amplification at C_T > 45, whereas the other NTC did not show any amplification. (B) Ethidium bromide stained gel electrophoresis in 1% agarose displays the semi-quantitative levels of DCP1B cDNA (expected and observed at ~850 bp) in post-mortem human brain tissues. No expression was detected in proband 1 (P1). The DCP1B cDNA levels were higher in the FC, compared to HC of proband 2 (P2) and proband 3 (P3). (A) gDNA amplicon band (expected and observed at ~6.9 kb) could be detected and no band in the NTC.

Figure 4

Box plot of the distribution of hsa-miR-138-1-5p expression levels in lymphoblastoid cell lines dependent on the rs9882688 genotype in 308 individuals of European descent. Horizontal lines represent median values, boxes represent interquartile ranges, and whiskers extend to 1.5× the interquartile range; values outside this range are depicted as circles. Carriers of the rs9882688 G allele showed a statistically significant (P = 0.000504) increase in hsa-miR-138-1-5p expression levels.