A Non-Synonymous HMGA2 Variant Decreases Height in Shetland Ponies and Other Small Horses - PubMed (original) (raw)

. 2015 Oct 16;10(10):e0140749.

doi: 10.1371/journal.pone.0140749. eCollection 2015.

Vidhya Jagannathan 2, Philippe Plattet 3, Markus Neuditschko 4, Heidi Signer-Hasler 5, Iris Bachmann 6, Alicja Pacholewska 7, Cord Drögemüller 2, Elisabeth Dietschi 2, Christine Flury 8, Stefan Rieder 4, Tosso Leeb 2

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A Non-Synonymous HMGA2 Variant Decreases Height in Shetland Ponies and Other Small Horses

Mirjam Frischknecht et al. PLoS One. 2015.

Abstract

The identification of quantitative trait loci (QTL) such as height and their underlying causative variants is still challenging and often requires large sample sizes. In humans hundreds of loci with small effects control the heritable portion of height variability. In domestic animals, typically only a few loci with comparatively large effects explain a major fraction of the heritability. We investigated height at withers in Shetland ponies and mapped a QTL to ECA 6 by genome-wide association (GWAS) using a small cohort of only 48 animals and the Illumina equine SNP70 BeadChip. Fine-mapping revealed a shared haplotype block of 793 kb in small Shetland ponies. The HMGA2 gene, known to be associated with height in horses and many other species, was located in the associated haplotype. After closing a gap in the equine reference genome we identified a non-synonymous variant in the first exon of HMGA2 in small Shetland ponies. The variant was predicted to affect the functionally important first AT-hook DNA binding domain of the HMGA2 protein (c.83G>A; p.G28E). We assessed the functional impact and found impaired DNA binding of a peptide with the mutant sequence in an electrophoretic mobility shift assay. This suggests that the HMGA2 variant also affects DNA binding in vivo and thus leads to reduced growth and a smaller stature in Shetland ponies. The identified HMGA2 variant also segregates in several other pony breeds but was not found in regular-sized horse breeds. We therefore conclude that we identified a quantitative trait nucleotide for height in horses.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1

Fig 1. Mapping of height QTL in Shetland ponies.

A: Manhattan plot of GWAS using 48 Shetland ponies and height at withers as a quantitative trait. B: Zoom of GWAS results on ECA 6. C: Haplotype analysis of the 19 smallest Shetland ponies. Each line represents one haplotype and the length of the lines represents the length of the shared haplotype block, which was defined based on a small Shetland pony that was homozygous for all significantly associated SNPs on ECA 6. D: NCBI MapViewer gene annotation in the critical interval (Annotation release 101, EquCab 2 genome assembly). The HMGA2 gene is currently not annotated in the NCBI MapViewer. Its position and orientation was determined by comparisons with the orthologous human genome segment.

Fig 2

Fig 2. Multiple species alignment of the first 37 amino acids of HMGA2.

The first AT-hook domain of HMGA2 is shaded in orange. Position 28 with the p.G28E variant in Shetland ponies is fully conserved in all analyzed vertebrate species including zebrafish The sequences were derived from the following accession numbers: Equus caballus, LN849001; Homo sapiens, NP_003474.1; Pan troglodytes, XP_009423258.1; Bos taurus, XP_002687694.2; Sus scrofa, XP_005664014.1; Mus musculus, NP_034571.1; Rattus norvegicus, NP_114459.1; Gallus gallus, NP_990332.1; Danio rerio, NP_997845.1.

Fig 3

Fig 3. A: Proportion of each genotype for the HMGA2:c.83G>A variant by breed.

The mutant allele occurred exclusively in small pony breeds.

Fig 4

Fig 4. Genotype-phenotype correlation.

The box plot displays the height at withers of Shetland ponies with the three different genotypes at the HMGA2:c.83G>A variant. The medians, which are indicated as solid horizontal lines in the box plot, were A/A: 84 cm, A/G: 98 cm, G/G: 104 cm. The mean heights per genotype were: A/A: 84 cm, A/G: 95 cm, G/G: 103 cm.

Fig 5

Fig 5. Electrophoretic mobility shift assay (EMSA).

We incubated a double-stranded oligonucleotide (dsDNA) with increasing concentrations of peptides containing either the wildtype (28Gly) or mutant (28Glu) sequence of the first AT-hook of the HMGA2 protein. With the wildtype sequence a partial shift of the dsDNA band could already be observed at the lowest concentration of peptide corresponding to a molar ratio of peptide to DNA of 1.4. At the highest concentration of the wildtype peptide, the majority of the DNA was in the DNA-protein complex. In contrast, with the mutant peptide the band shift was only observed at higher peptide concentrations and a smaller proportion of DNA was bound.

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