Systematic Functional Characterization of Candidate Causal Genes for Type 2 Diabetes Risk Variants

Soren K. Thomsen; Alessandro Ceroni; Martijn van de Bunt; Carla Burrows; Amy Barrett; Raphael Scharfmann; Daniel Ebner; Mark I. McCarthy; Anna L. Gloyn

doi:10.2337/db16-0361

Genetics/Genomes/Proteomics/Metabolomics

Systematic Functional Characterization of Candidate Causal Genes for Type 2 Diabetes Risk Variants

Soren K. Thomsen¹,
Alessandro Ceroni²,
Martijn van de Bunt¹,³,
Carla Burrows¹,
Amy Barrett¹,
Raphael Scharfmann⁴,
Daniel Ebner²,
Mark I. McCarthy¹,³,⁵ and
Anna L. Gloyn¹,³,⁵ ⇑

¹Oxford Centre for Diabetes, Endocrinology & Metabolism, Radcliffe Department of Medicine, University of Oxford, Oxford, U.K.
²Target Discovery Institute, Nuffield Department of Medicine, University of Oxford, Oxford, U.K.
³Wellcome Trust Centre for Human Genetics, Nuffield Department of Medicine, University of Oxford, U.K.
⁴INSERM U1016, Institut Cochin, Université Paris Descartes, Paris, France
⁵National Institute for Health Research Oxford Biomedical Research Centre, Churchill Hospital, Oxford, U.K.

Corresponding author: Anna L. Gloyn, anna.gloyn{at}drl.ox.ac.uk.

Diabetes 2016 Dec; 65(12): 3805-3811. https://doi.org/10.2337/db16-0361

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Figure 1
Comparing mean absolute effect sizes for MODY and non-MODY genes. Box plots of mean absolute effect sizes for MODY genes and non-MODY genes (excluding controls) across the five phenotypes measured. Effect sizes were calculated as described for Table 1, and the absolute values were then averaged for the two categories of genes. Among 14 identified MODY genes, 8 fulfilled criteria for inclusion in the screen: HNF4A, GCK, HNF1A, HNF1B, PAX4, INS, ABCC8, and KCNJ11. Tol, tolbutamide. Box plots show median and interquartile ranges for groups of n = 8 and 292 data points. ***q value <0.001 by Student t test (FDR-adjusted).
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Figure 2
Comparison of insulin secretion data for high and low glucose. Normalized insulin secretion responses under high glucose vs. low glucose, with selected hits annotated. The blue circle indicates the 95% confidence contour for NT control, and the orange circle indicates the 95% confidence contour for INS-positive controls. All measurements were normalized on a per-well basis to cell counts, and averages for each condition were then subsequently normalized to the mean of NT control. Data points are mean of n = 3 and shown as percentage of NT control.
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Figure 3
Insulin secretion data for selected genes in a follow-up validation experiment. Insulin secretion for ARL15 (A), ZMIZ1 (B), THADA (C), and HNF4A (D) (white bars) vs. NT (black bars) negative control under the indicated conditions. Measurements were processed as described for Fig. 2 and shown as percentage of NT control. Tol, tolbutamide. Data points are the mean of n = 6 for NT and n = 3 for other genes, and error bars are SEM. +q value <0.1, *q value <0.05, **q value <0.01, ***q value <0.001 by Student t test (FDR-adjusted).

Tables

Figures

Table 1

Effects of significant hits identified in a primary screen for β-cell dysfunction

Gene	Locus	Low glucose	High glucose	IBMX	Tolbutamide	Cell count
ABCC8	KCNJ11	48.2^*	24.0	26.7^*	1.8	−1.4
ADAMTS9	ADAMTS9	6.3	−4.8	2.8	−8.0	12.2^*
ADIPOQ	ST64GAL1	87.0^*	23.2	23.1	8.8	−7.3
ARL15	ARL15	−5.5	−25.9^*	−2.1	−15.5	−1.5
BCAR1	BCAR1	5.9	25.2	28.5^*	9.5	−7.0
BCL6	LPP	−20.7^*	−8.9	−1.0	−12.0	5.6
BMP8B	MACF1	7.8	16.5	9.5	26.9^*	−1.0
CCNT2	TMEM163	−32.6^*	1.8	4.7	5.0	−2.8
CDKAL1	CDKAL1	2.4	1.7	−10.6	−23.5^*	7.5
DGKQ	MAEA	3.3	17.5	20.4	33.6^*	−9.8
DMRTA2	FAF1	32.3^*	24.5	13.1	22.7	−0.3
ELAVL4	FAF1	−3.6	9.1	21.2	22.8	−11.4^*
ETV5	IGF2BP2	−12.4	−25.6^*	−10.9	−12.6	−2.5
FAH	ZFAND6	−23.1^*	−20.6^*	−16.9	−27.2	−2.3
FBXW7	TMEM154	45.2^*	9.6	8.7	11.7	9.9^*
GINS4	ANK1	−16.1	−14.2	−9.5	−17.0	8.7^*
GLIS3	GLIS3	−13.0	−10.6	6.4	−9.3	−10.3^*
HEYL	MACF1	29.2	29.2^*	0.4	20.0	−7.2
HMGA2	HMGA2	16.5	4.1	14.2	24.1^*	−1.2
HNF1A	HNF1A	21.7	38.3^*	23.1	25.5^*	5.2
HNF4A	HNF4A	36.7^*	66.9^*	74.9	92.9^*	−8.8
IGF2	DUSP8	−26.3^*	−10.4	1.2	0.1	−1.3
INS	DUSP8	−53.5^*	−44.8^*	−48.7	−38.7^*	0.9
KCNK17	KCNK16	−9.4	−17.3	−2.5	−1.2	9.8^*
KCTD15	PEPD	21.7	9.4	12.5	0.8	−10.9^*
KIF11	HHEX/IDE	45.4^*	35.0^*	26.9	55.4^*	−40.1^*
LINGO1	HMG20A	0	19.1	−12.4	−14.9	7.9^*
MFGE8	AP3S2	30.0^*	3.4	2.8	−1.7	−5.7
MIER3	ANKRD55	5.9	36.5^*	5.8	18.2	1.9
NDUFS4	ARL15	1.6	−4.9	3.9	−1.7	10.8^*
PABPC1L	HNF4A	−9.7	−12.3	−10.4	−28.8^*	−1.2
PHF23	SLC16A11	−25.6^*	−1.7	−5.9	3.5	−0.9
PLA2R1	RBMS1	8.6	1.6	10.8	1.6	9.1^*
PRDX3	GRK5	24.0	31.7^*	23.4	9.6	16.6^*
PTHLH	KLHDC5	−2.8	−6.5	−0.5	−25.0^*	−5.9
RND3	RND3	−8.7	−6.1	0	−3.0	−14.9^*
SLC2A4	SLC16A11	14.5	0	27.2^*	18.2	−1.6
SOCS7	HNF1B	3.9	−18.5^*	11.2	−14.7	−1.6
SPPL3	HNF1A	−11.9	−21.9^*	−6.1	−10.0	−10.8^*
STK38L	KLHDC5	15.2	40.9^*	4.7	25.2^*	−3.0
THADA	THADA	−1.9	6.5	27.5	24.8^*	−10.3
TLE1	TLE1	4.3	−5.0	−23.0^*	16.2	4.6
TM6SF2	CILP2	−22.6^*	−8.3	−0.8	−12.0	−8.7
UPF2	CDC123	7.5	−12.5	4.7	−24.9^*	−3.2
ZMIZ1	ZMIZ1	−29.5^*	−21.4^*	−19.8	−16.8	−15.2^*

The table lists effect sizes (% deviation from NT control) for each gene with a least one significant effect across the five phenotypes measured. All insulin secretion measurements were normalized on a per-well basis to cell counts, and the mean percentage deviations from NT control were then calculated for each condition. For cell counts, values were median-normalized for interplate differences, and the mean percentage deviations from NT control were calculated across conditions.
*q < 0.05 by Student t test (FDR-adjusted).