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Functional characterization of a haplotype in the AKT1 gene associated with glucose homeostasis and metabolic syndrome

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Abstract

A small 12-kb haplotype upstream of the AKT1 gene has been found to be associated with insulin resistance phenotypes. We sought to define the functional consequences of the three component polymorphic loci (rs1130214, rs10141867, rs33925946) on AKT1 and the upstream ZBTB42 gene. 5′ RACE analysis of AKT1 transcripts in human skeletal muscle biopsies showed the predominant promoter to be 2.5 kb upstream of exon 2, and distinct from those promoters previously reported in rat. We then studied the effect of each of the three haplotype polymorphisms in transcriptional reporter assays in muscle, bone, and fat cell culture models, and found that each modulated enhancer and repressor activity are in a cell-specific and differentiation-specific manner. Our results in promoter assays are consistent with the human phenotype data; we found an anabolic effect on muscle and bone with increased mRNA expression of AKT1, and catabolic effect on fat with decreased expression. To test the hypothesis that rs10141867 affects transcription levels of the novel zinc finger protein ZBTB42 in vivo, we developed the allele-specific expression assay using Taqman technology to test for allelic differences within heterozygotes. The allele containing the derived polymorphism (haplotype H2) showed a 1.75-fold increase in expression in human skeletal muscle. Our data show a particularly complex effect of the component polymorphisms of a single haplotype on cells and tissues, suggesting that the coordination of different tissue-specific effects may have driven selection for the H2 haplotype. In light of the recent abundance of SNP association studies, our approach can serve as a method for exploring the biological function of polymorphisms that show significant genotype/phenotype associations.

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Abbreviations

3T3-L1:

Murine fat cells

AKT1:

v-akt murine thymoma viral oncogene homolog 1

ASE:

Allele-specific expression assay

BMI:

Body mass index

C2C12:

Murine muscle cells

FAMUSS:

Functional SNPs associated with muscle size and strength

FOXO:

Forkhead box O

H1:

AKT1 haplotype H1

H2:

AKT1 haplotype H2

LPH:

Lactase-phlorizin hydrolase

MAF:

Minor allele frequency

MC3T3-E1:

Murine bone cells

MM:

Homozygote mutant

mTOR:

Mammalian target of rapamycin

mZBTB42:

Mouse ZBTB42

NN:

Homozygote normal

PDK:

PI-dependent kinases

PI3K:

Phosphatidylinositol 3 kinase

PKB:

Protein kinase B

PPARg:

Peroxisome proliferator-activated receptor gamma

RACE:

Rapid Amplification of cDNA Ends

RST:

Resistin

S6K1:

S6 kinase

SH2:

Src-homology 2

SNPs:

Single nucleotide polymorphisms

T2D:

Type 2 diabetes

UTR:

Untranslated region

WHO:

World Health Organization

YRI:

Yoruban HapMap population

ZBTB42:

Zinc finger and BTB domain containing 42

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Authors and Affiliations

  1. Department of Integrative Systems Biology, Research Center for Genetic Medicine, Children’s National Medical Center, 111 Michigan Avenue NW, Washington, DC, 20010, USA

    Brennan T. Harmon, Stephanie A. Devaney, Heather Gordish-Dressman, Erica K. Reeves, Po Zhao, Joseph M. Devaney & Eric P. Hoffman

  2. Department of Integrative Systems Biology, George Washington University School of Medicine and Health Sciences, Washington, DC, USA

    Eric P. Hoffman

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  1. Brennan T. Harmon
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  2. Stephanie A. Devaney
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  3. Heather Gordish-Dressman
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  4. Erica K. Reeves
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  5. Po Zhao
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  6. Joseph M. Devaney
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  7. Eric P. Hoffman
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Correspondence to Eric P. Hoffman.

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Harmon, B.T., Devaney, S.A., Gordish-Dressman, H. et al. Functional characterization of a haplotype in the AKT1 gene associated with glucose homeostasis and metabolic syndrome. Hum Genet 128, 635–645 (2010). https://doi.org/10.1007/s00439-010-0891-7

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