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Identification of microRNAs associated with allergic airway disease using a genetically diverse mouse population.

Rutledge H, Baran-Gale J, de Villena FP, Chesler EJ, Churchill GA, Sethupathy P, Kelada SN - BMC Genomics (2015)

Bottom Line: Among CC founders, 92 miRNAs were differentially expressed.Analysis of a dataset from human keratinocytes transfected with a miR-31 inhibitor revealed two target genes in common with miR-31 targets correlated with neutrophils, namely Oxsr1 and Nsf. miRNA expression in the allergically inflamed murine lung is regulated by genetic loci that are smaller in effect size compared to mRNA eQTL and often act in trans.We identified three miRNAs, miR-497, miR-351 and miR-31, that are candidate master regulators of genes associated with neutrophil recruitment.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, University of North Carolina, 120 Mason Farm Road, Chapel Hill, NC, 27599, USA. hrutledg@email.unc.edu.

ABSTRACT

Background: Allergic airway diseases (AADs) such as asthma are characterized in part by granulocytic airway inflammation. The gene regulatory networks that govern granulocyte recruitment are poorly understood, but evidence is accruing that microRNAs (miRNAs) play an important role. To identify miRNAs that may underlie AADs, we used two complementary approaches that leveraged the genotypic and phenotypic diversity of the Collaborative Cross (CC) mouse population. In the first approach, we sought to identify miRNA expression quantitative trait loci (eQTL) that overlap QTL for AAD-related phenotypes. Specifically, CC founder strains and incipient lines of the CC were sensitized and challenged with house dust mite allergen followed by measurement of granulocyte recruitment to the lung. Total lung RNA was isolated and miRNA was measured using arrays for CC founders and qRT-PCR for incipient CC lines.

Results: Among CC founders, 92 miRNAs were differentially expressed. We measured the expression of 40 of the most highly expressed of these 92 miRNAs in the incipient lines of the CC and identified 18 eQTL corresponding to 14 different miRNAs. Surprisingly, half of these eQTL were distal to the corresponding miRNAs, and even on different chromosomes. One of the largest-effect local miRNA eQTL was for miR-342-3p, for which we identified putative causal variants by bioinformatic analysis of the effects of single nucleotide polymorphisms on RNA structure. None of the miRNA eQTL co-localized with QTL for eosinophil or neutrophil recruitment. In the second approach, we constructed putative miRNA/mRNA regulatory networks and identified three miRNAs (miR-497, miR-351 and miR-31) as candidate master regulators of genes associated with neutrophil recruitment. Analysis of a dataset from human keratinocytes transfected with a miR-31 inhibitor revealed two target genes in common with miR-31 targets correlated with neutrophils, namely Oxsr1 and Nsf.

Conclusions: miRNA expression in the allergically inflamed murine lung is regulated by genetic loci that are smaller in effect size compared to mRNA eQTL and often act in trans. Thus our results indicate that the genetic architecture of miRNA expression is different from mRNA expression. We identified three miRNAs, miR-497, miR-351 and miR-31, that are candidate master regulators of genes associated with neutrophil recruitment. Because miR-31 is expressed in airway epithelia and is predicted to target genes with known links to neutrophilic inflammation, we suggest that miR-31 is a potentially novel regulator of airway inflammation.

No MeSH data available.


Related in: MedlinePlus

The miR-342-3p eQTL. a. Expression of miR-342-3p in CC founders (a) and preCC mice (b) as a function of founder haplotype at the eQTL on chromosome 12. CC founder miRNA expression was measured by microarray while preCC miRNA expression was measured by qRT-PCR and data for the latter are presented as −1*DeltaCq (normalized to miR-17*). c. Phylogeny of CC founder strains based on SNP data for the region on Chr 12 containing the miR-342-3p eQTL. Bootstrap values were greater than or equal to 96 (out of 100) for each branch of tree.The red line denotes the branch that contains the putative causal variant
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Fig4: The miR-342-3p eQTL. a. Expression of miR-342-3p in CC founders (a) and preCC mice (b) as a function of founder haplotype at the eQTL on chromosome 12. CC founder miRNA expression was measured by microarray while preCC miRNA expression was measured by qRT-PCR and data for the latter are presented as −1*DeltaCq (normalized to miR-17*). c. Phylogeny of CC founder strains based on SNP data for the region on Chr 12 containing the miR-342-3p eQTL. Bootstrap values were greater than or equal to 96 (out of 100) for each branch of tree.The red line denotes the branch that contains the putative causal variant

Mentions: We categorized the miRNA eQTL in terms of their effect size and location in the genome. Two eQTL, for miR-489 and miR-342-3p, stood out in terms of effect size. Not surprisingly, expression of these two miRNAs also showed the highest heritability in CC founder lines (Additional file 5: Table S2). We confirmed that for both miRNAs the expression levels among preCC mice with a specific CC founder haplotype at the eQTL were concordant with expression levels in the CC founder strain (Figs. 3 and 4). We calculated the narrow sense heritability (h2) to be 0.60 and 0.84 for the miR-489 and miR-342-3p eQTL, respectively. Taken together, these results indicate that cis-regulatory elements at the respective eQTL are the primary determinants of miR-489 and miR-342-3p expression in the lung. However, we note that miR-489 expression in CC founders suggests three distinct groups (high expression group composed of NOD/ShiLtJ and WSB/EiJ; medium expression group composed of A/J, C57BL/6J, 129S1SvImJ, and NZO/H1LtJ; and low expression group composed of CAST/EiJ and PWK/PhJ), but the allele effects for the eQTL on chromosome (Chr) 6 indicates that this eQTL only explains the differences between CAST/EiJ and PWK/PhJ vs. the other six CC founders. Thus it appears that one or more additional loci likely contribute to miR-489 expression; but we did not identify other eQTL (local or distant) using genome scans in which we conditioned on the Chr 6 eQTL.Fig. 3


Identification of microRNAs associated with allergic airway disease using a genetically diverse mouse population.

Rutledge H, Baran-Gale J, de Villena FP, Chesler EJ, Churchill GA, Sethupathy P, Kelada SN - BMC Genomics (2015)

The miR-342-3p eQTL. a. Expression of miR-342-3p in CC founders (a) and preCC mice (b) as a function of founder haplotype at the eQTL on chromosome 12. CC founder miRNA expression was measured by microarray while preCC miRNA expression was measured by qRT-PCR and data for the latter are presented as −1*DeltaCq (normalized to miR-17*). c. Phylogeny of CC founder strains based on SNP data for the region on Chr 12 containing the miR-342-3p eQTL. Bootstrap values were greater than or equal to 96 (out of 100) for each branch of tree.The red line denotes the branch that contains the putative causal variant
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4548451&req=5

Fig4: The miR-342-3p eQTL. a. Expression of miR-342-3p in CC founders (a) and preCC mice (b) as a function of founder haplotype at the eQTL on chromosome 12. CC founder miRNA expression was measured by microarray while preCC miRNA expression was measured by qRT-PCR and data for the latter are presented as −1*DeltaCq (normalized to miR-17*). c. Phylogeny of CC founder strains based on SNP data for the region on Chr 12 containing the miR-342-3p eQTL. Bootstrap values were greater than or equal to 96 (out of 100) for each branch of tree.The red line denotes the branch that contains the putative causal variant
Mentions: We categorized the miRNA eQTL in terms of their effect size and location in the genome. Two eQTL, for miR-489 and miR-342-3p, stood out in terms of effect size. Not surprisingly, expression of these two miRNAs also showed the highest heritability in CC founder lines (Additional file 5: Table S2). We confirmed that for both miRNAs the expression levels among preCC mice with a specific CC founder haplotype at the eQTL were concordant with expression levels in the CC founder strain (Figs. 3 and 4). We calculated the narrow sense heritability (h2) to be 0.60 and 0.84 for the miR-489 and miR-342-3p eQTL, respectively. Taken together, these results indicate that cis-regulatory elements at the respective eQTL are the primary determinants of miR-489 and miR-342-3p expression in the lung. However, we note that miR-489 expression in CC founders suggests three distinct groups (high expression group composed of NOD/ShiLtJ and WSB/EiJ; medium expression group composed of A/J, C57BL/6J, 129S1SvImJ, and NZO/H1LtJ; and low expression group composed of CAST/EiJ and PWK/PhJ), but the allele effects for the eQTL on chromosome (Chr) 6 indicates that this eQTL only explains the differences between CAST/EiJ and PWK/PhJ vs. the other six CC founders. Thus it appears that one or more additional loci likely contribute to miR-489 expression; but we did not identify other eQTL (local or distant) using genome scans in which we conditioned on the Chr 6 eQTL.Fig. 3

Bottom Line: Among CC founders, 92 miRNAs were differentially expressed.Analysis of a dataset from human keratinocytes transfected with a miR-31 inhibitor revealed two target genes in common with miR-31 targets correlated with neutrophils, namely Oxsr1 and Nsf. miRNA expression in the allergically inflamed murine lung is regulated by genetic loci that are smaller in effect size compared to mRNA eQTL and often act in trans.We identified three miRNAs, miR-497, miR-351 and miR-31, that are candidate master regulators of genes associated with neutrophil recruitment.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, University of North Carolina, 120 Mason Farm Road, Chapel Hill, NC, 27599, USA. hrutledg@email.unc.edu.

ABSTRACT

Background: Allergic airway diseases (AADs) such as asthma are characterized in part by granulocytic airway inflammation. The gene regulatory networks that govern granulocyte recruitment are poorly understood, but evidence is accruing that microRNAs (miRNAs) play an important role. To identify miRNAs that may underlie AADs, we used two complementary approaches that leveraged the genotypic and phenotypic diversity of the Collaborative Cross (CC) mouse population. In the first approach, we sought to identify miRNA expression quantitative trait loci (eQTL) that overlap QTL for AAD-related phenotypes. Specifically, CC founder strains and incipient lines of the CC were sensitized and challenged with house dust mite allergen followed by measurement of granulocyte recruitment to the lung. Total lung RNA was isolated and miRNA was measured using arrays for CC founders and qRT-PCR for incipient CC lines.

Results: Among CC founders, 92 miRNAs were differentially expressed. We measured the expression of 40 of the most highly expressed of these 92 miRNAs in the incipient lines of the CC and identified 18 eQTL corresponding to 14 different miRNAs. Surprisingly, half of these eQTL were distal to the corresponding miRNAs, and even on different chromosomes. One of the largest-effect local miRNA eQTL was for miR-342-3p, for which we identified putative causal variants by bioinformatic analysis of the effects of single nucleotide polymorphisms on RNA structure. None of the miRNA eQTL co-localized with QTL for eosinophil or neutrophil recruitment. In the second approach, we constructed putative miRNA/mRNA regulatory networks and identified three miRNAs (miR-497, miR-351 and miR-31) as candidate master regulators of genes associated with neutrophil recruitment. Analysis of a dataset from human keratinocytes transfected with a miR-31 inhibitor revealed two target genes in common with miR-31 targets correlated with neutrophils, namely Oxsr1 and Nsf.

Conclusions: miRNA expression in the allergically inflamed murine lung is regulated by genetic loci that are smaller in effect size compared to mRNA eQTL and often act in trans. Thus our results indicate that the genetic architecture of miRNA expression is different from mRNA expression. We identified three miRNAs, miR-497, miR-351 and miR-31, that are candidate master regulators of genes associated with neutrophil recruitment. Because miR-31 is expressed in airway epithelia and is predicted to target genes with known links to neutrophilic inflammation, we suggest that miR-31 is a potentially novel regulator of airway inflammation.

No MeSH data available.


Related in: MedlinePlus