Trisomy-21 gene dosage over-expression of miRNAs results in the haploinsufficiency of specific target proteins

Figures & data

Figure 1 Relative expression of Hsa21-derived miRNAs in human tissues. Mature human let-7c, miR-99a, miR-125b, miR-155 and miR-802 were quantified utilizing TaqMan microRNA assay kits specific for each Hsa21-derived miRNA (Applied Biosystems, Foster City, CA) as previously described.40,42 The expression values were normalized to RNU48 for each tissue. Relative gene expression was calculated as 2^{-(CTmiR-155-CT-RNU48)}. The relative abundance of each Hsa-21 derived miRNA is shown as a percentage of total combined expression of Hsa-21 derived miRNAs for each tissue investigated.

Figure 2 Hsa21-derived miRNAs are overexpressed in human DS pre-frontal cortex brain specimens. Mature Hsa21-derived miRNAs were quantified utilizing RT-PCR as previously described by our laboratory40,42,85,86 using total RNA isolated from human fetal (18–22 weeks of gestation), child (1–8 years), adolescent (9–19 years old) and adult (20–50 years old) prefrontal cortex specimens from control and DS (age- and sex-matched, n = 3–5) samples. Gene expression was calculated relative to 18S rRNA and the data are expressed as percent over control (variation between control samples was never greater than 5%), which was assigned a value of 100%. The error bars represent the average ± S.E. of triplicate samples repeated in at least three independent experiments (*p < 0.01 DS vs. control).

Figure 3 The human AT₁R +1166 A/C SNP occurs in the miR-155-binding site. (A) Complementarity between miR-155 and the hAT₁R 3′-UTR site targeted (70–90 bp downstream from the human AT₁R stop codon). The +1166 A/C SNP corresponds to the nucleotide 86 bp downstream from the human AT₁R stop codon (shown in bold print). The binding of miR-155 to the hAT₁R 3′-UTR target site fulfills the requirement of a 7 bp seed sequence of complementarity at the miRNA 5′ end when the +1166 A-allele is expressed. (B) Complementarity between miR-155 and the human AT₁R 3′-UTR harboring the +1166 C-allele. If the +1166 C-allele is expressed, the seed sequence requirement would not be met and, as a consequence, it would be expected that human AT₁R expression would be elevated.

Table 1 Targetscan algorithm-predicted Hsa21-derived putative miRNA/mRNA targets

Hsa-21-derived miRNA	Potential human targets*	Number of targets containing at least two potential binding sites	Number of targets containing at least three potential binding sites	Number of targets containing at least four potential binding sites	Number of targets containing at least five potential binding sites
let-7c	819
miR-99a	40
miR-125b-2	604
miR-155	281
miR-802	232
Total	1695	541	194	33	0

* Source: TargetScan (April, 2009). The number of potential targets is dramatically dependent upon the algorithm utilized.

Kuhn DE, Nuovo GJ, Martin MM, Malana GE, Pleister AP, Jiang J, et al. Human chromosome 21-derived miRNAs are overexpressed in down syndrome brains and hearts. Biochem Biophys Res Commun 2008; 370:473 - 477

 PubMed Web of Science ®Google Scholar

Kuhn DE, Nuovo GJ, Terry A Jr, Martin MM, Malana GE, Sansom SE, et al. Chromosome 21-derived mirnas provide an etiological basis for aberrant protein expression in human down syndrome brains. J Biol Chem 2010; 285:1529 - 1543

 PubMed Web of Science ®Google Scholar

Martin MM, Lee EJ, Buckenberger JA, Schmittgen TD, Elton TS. MicroRNA-155 regulates human angiotensin II type 1 receptor expression in fibroblasts. J Biol Chem 2006; 281:18277 - 84

 PubMed Web of Science ®Google Scholar

Martin MM, Buckenberger JA, Jiang J, Malana GE, Nuovo GJ, Chotani M, et al. The human angiotensin II type 1 receptor +1166 A/C polymorphism attenuates microrna-155 binding. J Biol Chem 2007; 282:24262 - 24269

 PubMed Web of Science ®Google Scholar