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International Textbook of Diabetes Mellitus, 4th Ed., Excerpt #141: The Genetics of Type 2 Diabetes Part 3

Sep 4, 2018

Identification of T2DM affecting genetic variants

Linkage studies

The methods used to map disease-causing variation have evolved rapidly in the last decades thanks to technical advances in genotyping methods (Figure 26.2). Originally, disease-causing loci were identified primarily by linkage analysis, utilizing the long stretches of linkage in affected families. By genotyping 400–500 genetic markers, disease loci can be mapped on a genome-wide level without any prior hypothesis about which genes are involved. Finding that affected family members share a certain marker that is identical by descent (i.e., identical because it was inherited from the same parent) more often than expected by chance is evidence that a disease-causing variant is in linkage with that marker. This strategy has been very successful in mapping genetic diseases like MODY that have a strong penetrance and a known mode of inheritance. The first MODY locus, MODY-2, was mapped to the glucokinase (GK) locus on chromosome 7 in 1992 [10]. It was soon followed by the HNF1A (MODY-3) and HNF4A (MODY-1) loci [11,12]. The HNF homeobox genes are transcription factors expressed in liver and β-cells that are necessary for proper regulation of insulin secretion. Common variants in both these loci have since been shown to affect risk of T2DM [13].

Linkage analysis has, however, been less useful for identifying genes causing complex diseases. Even though great efforts have been put into linkage studies of T2DM, only two genes can be claimed to have been identified using this strategy. The first T2DM gene mapped by linkage analysis was CAPN10 on chromosome 10, encoding calpain 10, a cysteine protease with largely unknown functions in glucose metabolism [14]. Despite a number of negative replication studies, several meta-analyses have shown consistent association with T2DM[15,16]. Nevertheless, none of the large genome-wide association studies (GWAS) have identified CAPN10 as being associated with T2DM.

The second locus was first mapped to a 10.5Mb region on chromosome 10q and was later fine-mapped in the Icelandic population to an intronic variant in the TCF7L2 gene contributing to, but not fully explaining, the original linkage [17–19]. This association has since been confirmed in African, Asian, and European populations making it the best replicated genetic association with T2DM to date, conferring a relative risk of ∼1.4 [20].

Association studies on candidate genes

A popular hypothesis about the genetic architecture of complex diseases such as T2DM, the common disease/common variant hypothesis, suggests that common disorders are caused by aggregation of common risk alleles [21,22]. This model has been the basis of a revolution in complex genetics by stimulating the development of tools for genetic association studies. Association studies utilize the very short LD stretches in unrelated individuals to map risk variants in populations instead of families. The advantages of association lie in the larger number of individuals that can be collected for each study as well as the much higher resolution of the mapping. One disadvantage is the huge number of markers needed to perform mapping on a genome-wide level and association studies were therefore originally performed on small regions known to harbor candidate genes.

The first gene reproducibly associated with T2DM was PPARG, encoding the nuclear receptor  PPAR-γ [23]. The PPAR-γ receptor is a molecular target for thiazolidinedione compounds, a class of insulin-sensitizing drugs used to treat T2DM, making it a very compelling candidate gene. The transcript expressed in adipose tissue has an extra exon B and a substitution of a proline for alanine at position 12 of this protein, which is seen in about 15% of the European population. This variant, has been shown to be associated with increased transcriptional activity, increased insulin sensitivity, and protection against T2DM [23].

The ADRA2A (adrenergic receptor alpha 2) locus was recently identified as a T2DM risk locus after first having been positionally mapped in congenic GK rats where it was associated with impaired insulin granule docking and reduced β-cell exocytosis [24]. Human carriers of the ADRA2A risk variant (rs553668) have reduced fasting insulin and decreased insulin secretion as a consequence of increased expression of the ADRA2 receptor in pancreatic islets. It is well known that epinephrine excess can suppress insulin secretion and cause diabetes.

Genome-wide association studies (GWAS)

Rapid improvement in high throughput technology for SNP genotyping, allowing simultaneous genotyping of hundreds of thousands of SNPs, has opened new possibilities for association studies. The HapMap project provided another important tool, showing that genotyping of approximately 500,000 SNPs is enough to cover about 75% of the common variants (minor allele frequency >5%) in the genome. Several GWAS for diabetes were published in 2007, coined “Breakthrough of the Year” by Science magazine.The first was a GWAS on early-onset T2DM that was published in February 2007 reporting two new diabetes loci: HHEX and SLC30A8 [25]. A few months later it was followed by four European case-control studies including patients with classical T2DM [26–29]. Three of the studies shared results prior to publication and only considered positive results that were replicated in all studies. This resulted in the identification of two new diabetes loci, CDKN2A/2B and IGF2BP2, in addition to confirming previously known loci. CDKAL1 (CDK5 regulatory subunit associated protein 1-like 1) was independently identified as a new T2DM locus in all four studies.

About the same time FTO was identified as a major susceptibility locus for obesity, and therefore indirectly also for T2DM [30,31]. Since the effect of FTO on diabetes is through obesity it was not detected in the GWAS studies of T2DM that were matched on BMI.

The first GWA studies on T2DM in non-European populations were published in 2008 using a multistage approach [32,33]. Both studies identified KCNQ1, which encodes the pore-forming alpha subunit of the IKsK+ channel (or voltage-gated potassium channel), and showed that the gene is expressed in the pancreas. Later a SNP in KCNQ1 was shown to have distorted parent-of-origin transmission in an Icelandic study, the risk allele having a much stronger effect when transmitted from the mother than from the father [34].

The first wave of GWAS was followed by a second wave combining existing or new GWAS into meta-analyses of >50,000 individuals [13,35]. A prerequisite for this was that many research groups could work together in consortia like DIAGRAM (DIAbetes Genetics Replication and Meta-analysis) consortium and MAGIC (Meta-Analyses of Glucose- and Insulin-related traits consortium).These very large studies have identified several new loci and confirmed the effect of many previously identified. This has also been facilitated by the 1000 Genomes Project, another international collaboration to produce a public catalog of human genetic variation, including SNPs and structural variants,which now allows imputation and analysis of more than 30 million common and low-frequency variants.

Recently, a number of GWAS and meta-analysis studies have also been performed in non-European cohorts, adding several new loci to the list of genome-wide significant associations [35–45]. Interestingly it seems like most associations found in one ethnic group  also show some evidence of association in populations with other ethnicities. In total, GWAS have provided more than 75 loci for T2DM (Table 26.1) as well as numerous loci for glucose- or insulin-related traits and more are likely to come.

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