Abstract
Resolving the underlying functional mechanism to a given genetic association has proven extremely challenging. However, the strongest associated type 2 diabetes (T2D) locus reported to date, TCF7L2, presents an opportunity for translational analyses, as many studies in multiple ethnicities strongly point to SNP rs7903146 in intron 3 as being the causal variant within this gene. We carried out oligo pull-down combined with mass spectrophotometry (MS) to elucidate the specific transcriptional machinery across this SNP using protein extracts from HCT116 cells. We observed that poly (ADP-ribose) polymerase 1 (PARP-1) is by far the most abundant binding factor. Pursuing the possibility of a feedback mechanism, we observed that PARP-1, along with the next most abundant binding proteins, DNA topoisomerase I and ATP-dependent RNA helicase A, dimerize with the TCF7L2 protein and with each other. We uncovered further evidence of a feedback mechanism using a luciferase reporter approach, including observing expression differences between alleles for rs7903146. We also found that there was an allelic difference in the MS results for proteins with less abundant binding, namely X-ray repair cross-complementing 5 and RPA/p70. Our results point to a protein complex binding across rs7903146 within TCF7L2 and suggests a possible mechanism by which this locus confers its T2D risk.
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Acknowledgements
A summary of the findings can be found at the TCF7L2 gene variant database located at: http://databases.lovd.nl/shared/variants/0000023424. This work was supported by the Institutional Development Funds from the Children’s Hospital of Philadelphia and an award from the Pennsylvania Department of Health. The University of Pennsylvania Proteomics Core is supported by NIH grants P30 CA016520 (Abramson Cancer Center) and P30 ES013508-04 (Center of Excellence in Environmental Toxicology).
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QX, SD, CXY, MEJ and SFAG researched data. QX and SFAG wrote manuscript. SD, CXY and MEJ reviewed/edited manuscript.
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Xia, Q., Deliard, S., Yuan, CX. et al. Characterization of the transcriptional machinery bound across the widely presumed type 2 diabetes causal variant, rs7903146, within TCF7L2. Eur J Hum Genet 23, 103–109 (2015). https://doi.org/10.1038/ejhg.2014.48
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DOI: https://doi.org/10.1038/ejhg.2014.48
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