Kyuyoung Song

Interaction of human Ku70 with TRF2

Ku, a heterodimer of 70‐ and 80‐kDa subunits, plays a general role in the metabolism of DNA ends in eukaryotic cells, including double‐strand DNA break repair, V(D)J recombination, and maintenance of telomeres. We have utilized the yeast two‐hybrid system to identify Ku70‐interacting proteins other than Ku80. Two reactive clones were found to encode the dimerization domain of TRF2, a mammalian telomeric protein that binds to duplex TTAGGG repeats at chromosome ends. This interaction was confirmed using bacterial fusion proteins and co‐immunoprecipitations from eukaryotic cells overexpressing TRF2. The transfected TFR2 colocalized with Ku70.

A common missense variant in NUDT15 confers susceptibility to thiopurine-induced leukopenia

Thiopurine therapy, commonly used in autoimmune conditions, can be complicated by life-threatening leukopenia. This leukopenia is associated with genetic variation in TPMT (encoding thiopurine S-methyltransferase). Despite a lower frequency of TPMT mutations in Asians, the incidence of thiopurine-induced leukopenia is higher in Asians than in individuals of European descent. Here we performed an Immunochip-based 2-stage association study in 978 Korean subjects with Crohns disease treated with thiopurines. We identified a nonsynonymous SNP in NUDT15 (encoding p.Arg139Cys) that was strongly associated with thiopurine-induced early leukopenia (odds ratio (OR) = 35.6; Pcombined = 4.88 × 10−94). In Koreans, this variant demonstrated sensitivity and specificity of 89.4% and 93.2%, respectively, for thiopurine-induced early leukopenia (in comparison to 12.1% and 97.6% for TPMT variants). Although rare, this SNP was also strongly associated with thiopurine-induced leukopenia in subjects with inflammatory bowel disease of European descent (OR = 9.50; P = 4.64 × 10−4). Thus, NUDT15 is a pharmacogenetic determinant for thiopurine-induced leukopenia in diverse populations.

Wnt5a Induces Endothelial Inflammation via β-Catenin–Independent Signaling

Wnt signaling has been implicated in certain inflammatory diseases. However, the biological role in the inflammatory regulation remains to be characterized. We investigated the regulation by Wnt signaling in endothelial cells, which are active participants and regulators of inflammation. Wnt5a induces cyclooxygenase-2 expression and enhances inflammatory cytokines rapidly, whereas Wnt3a shows limited effects, suggesting a role for β-catenin–independent Wnt signaling in the inflammatory endothelial activation. Pulse-like treatment of Wnt5a induces cyclooxygenase-2 more efficiently than continuous treatment. Wnt5a and TNF-α regulate subsets of cytokines overlapping, only partially, with each other. Calcium ionophore enhances endothelial inflammation similarly, whereas calcium chelators and protein kinase C inhibitor block Wnt5a-induced activation, suggesting a role for the Wnt/Ca2+/protein kinase C pathway in endothelial inflammatory regulation. Wnt5a activates RelA nuclear translocation and DNA binding. Activated blood vessels, histiocytes, and synoviocytes express Wnt5a in atherosclerosis and rheumatoid arthritis but not in normal tissue, supporting the role of Wnt5a as an inflammatory mediator in vivo. Our data suggest that endothelial inflammation is regulated by a dual system consisting of β-catenin–independent Wnt signaling and TNF-α–mediated signaling.

Genome-wide association study of Crohn's disease in Koreans revealed three new susceptibility loci and common attributes of genetic susceptibility across ethnic populations

Objective Crohns disease (CD) is an intractable inflammatory bowel disease (IBD) of unknown cause. Recent meta-analysis of the genome-wide association studies (GWAS) and Immunochip data identified 163 susceptibility loci to IBD in Caucasians, however there are limited studies in other populations. Methods We performed a GWAS and two validation studies in the Korean population comprising a total of 2311 patients with CD and 2442 controls. Results We confirmed four previously reported loci: TNFSF15, IL23R, the major histocompatibility complex region, and the RNASET2-FGFR1OP-CCR6 region. We identified three new susceptibility loci at genome-wide significance: rs6856616 at 4p14 (OR=1.43, combined p=3.60×10−14), rs11195128 at 10q25 (OR=1.42, combined p=1.55×10−10) and rs11235667 at 11q13 (OR=1.46, combined p=7.15×10−9), implicating ATG16L2 and/or FCHSD2 as novel susceptibility genes for CD. Further analysis of the 11q13 locus revealed a non-synonymous single nucleotide polymorphism (SNP) (R220W/rs11235604) in the evolutionarily conserved region of ATG16L2 with stronger association (OR=1.61, combined p=2.44×10−12) than rs11235667, suggesting ATG16L2 as a novel susceptibility gene for CD and rs11235604 to be a potential causal variant of the association. Two of the three SNPs (rs6856616 (p=0.00024) and rs11195128 (p=5.32×10−5)) showed consistent patterns of association in the International IBD Genetics Consortium dataset. Together, the novel and replicated loci accounted for 5.31% of the total genetic variance for CD risk in Koreans. Conclusions Our study provides new biological insight to CD and supports the complementary value of genetic studies in different populations.

Association of TNFSF15 with Crohn's disease in Koreans.

OBJECTIVES:A recent genomewide association study from a Japanese population identified tumor necrosis factor superfamily member 15 (TNFSF15) as an inflammatory bowel disease gene. Previous studies have shown that expression of TNFSF15 was upregulated in macrophages and lymphocytes of the intestinal lamina propria of Crohns disease (CD) patients. Here, we have tested four single nucleotide polymorphisms (SNPs) of TNFSF15 in Korean patients to determine whether the gene is associated with susceptibility to CD in a closely related population.METHODS:Four SNPs across TNFSF15 were genotyped in 380 patients with CD and 380 healthy controls.RESULTS:Carriers of three polymorphisms, including rs3810936, rs6478108, and rs7848647, showed statistically significant association with CD (adjusted OR [aOR] 2.81, 95% confidence interval [CI] 1.94–4.07, P= 4.4 × 10−8; aOR 3.49, 95% CI 2.42–5.04, P= 2.7 × 10−11; and aOR 3.49, 95% CI 2.42–5.03, P= 2.2 × 10−11, respectively). Following haplotype analysis, homozygotes carrying two copies of the haplotype consisting of the risk alleles of those three SNPs showed statistically significant association with CD (aOR 5.39, 95% CI 3.19–9.10, P= 3.07 × 10−10).CONCLUSIONS:Our data support the hypothesis that the TNFSF15 genotypes play an important role in the pathogenesis of CD in Koreans.

Human Ku70 Interacts with Heterochromatin Protein 1α

Ku is involved in the metabolism of DNA ends, DNA repair, and the maintenance of telomeres. It consists of a heterodimer of 70- and 80-kDa subunits. Recently we have demonstrated that Ku70 interacted with TRF2, a mammalian telomere-binding protein. Using the same yeast two-hybrid screening system, we now show that Ku70 also interacts with heterochromatin protein 1α (HP1α), a protein known to be associated with telomeres as well as heterochromatin. HP1 is a suppressor of the position effect variegation inDrosophila and acts as a transcriptional suppressor in mammalian cells. The interaction with Ku70 in the two-hybrid system was confirmed by a glutathione S-transferase pull-down study using bacterial recombinant proteins in vitro. The interaction was also reproduced in vivo in HeLa cells, where endogenous Ku70 coimmunoprecipitated with HP1α. This interaction was more effective in acidic pH and weakened considerably as the pH of the reaction buffer was elevated up to 7.5. Ku80 did not interact with HP1α directly. The interaction domains of Ku70 and HP1α included the Leu-Ser repeat (amino acids 200–385) and the chromo shadow domain, respectively. Ku70 was largely colocalized with transfected HP1α but not with a C-terminal deletion mutant, HP1αΔ C. In contrast to HP1α, Ku70 did not repress transcriptional activity of the reporter gene when tethered to DNA after transfection to mammalian cells. The implication of this interaction is discussed.

Contribution of IL23R but not ATG16L1 to Crohn's disease susceptibility in Koreans

Background: Recent genome‐wide association studies in Caucasian populations identified IL23R and ATG16L1 as susceptibility genes to Crohns disease (CD). We tested 5 IL23R single nucleotide polymorphisms (SNPs) and 12 ATG16L1 SNPs in Korean patients to determine whether these genes are associated with susceptibility to CD in a non‐Caucasian population. Methods: We analyzed 5 IL23R SNPs and 12 ATG16L1 SNPs in 380 patients with CD and 380 healthy controls. Results: Two IL23R gene variants, an intronic SNP rs1004819 and intergenic SNP rs1495465, showed significant associations with CD; the adjusted odds ratio (aOR) for rs1004819 was 1.822 (95% confidence interval [CI] = 1.164–2.852, P = 0.009) and aOR for rs1495965 was 1.650 (95% CI = 1.102–2.471, P = 0.015). The genotype–phenotype analysis showed subphenotype specificity to stricturing and penetrating behaviors. On the other hand, none of the 12 ATG16L1 SNPs showed any positive association with CD in Koreans. The contribution of IL23R variants in Korean CD patients overall is low in comparison with studies of Caucasian. Conclusions: Our data in Koreans support the previous Caucasian reports of an association of the IL23R gene with CD. (Inflamm Bowel Dis 2009)

Lack of association between pro-inflammatory genotypes of the interleukin-1 (IL-1B -31 C/+ and IL-1RN *2/*2) and gastric cancer/duodenal ulcer in Korean population.

IL-1beta is a pro-inflammatory cytokine with multiple biological effects and is a potent inhibitor of gastric acid secretion, and IL-1RN has been shown to be associated with enhanced IL-1beta production in vitro. Recently, it was reported that the pro-inflammatory genotypes, IL-1B -31 C/+ and IL-1RN *2/*2, were associated with an increased risk of gastric cancer in a Caucasian population. We tested the association between the polymorphisms and 190 gastric cancer, 117 duodenal ulcer, and 172 healthy subjects as controls in the Korean population. The allele frequency of IL-1B -31 C was more prevalent in Korean (51%) than in Caucasian (30%), while the frequency of IL-1RN *2 allele was less in Korean (6%) than in Caucasian (27%). Using the IL-1B TT genotype as a reference group, the CC genotype was not associated with an increased risk of gastric cancer or duodenal ulcer in the Korean population (odds ratios (OR)=0.90, 95% confidence interval (CI)=0.50-1.64; OR=0.72, 95% CI=0.36-1.46, respectively). Similarly, IL-1RN*2 was not a risk genotype for either gastric cancer or duodenal ulcer. No association was recognized on the haplotype analysis of the two genes, either. Our results did not support the previous report that IL-1B -31 C/IL-1RN*2 polymorphisms were associated with an increased risk of gastric cancer. The lack of association with duodenal ulcer also suggested that the polymorphisms were not directly related to the acid-secreting capability.

Long-term effects of a structured intensive diabetes education programme (SIDEP) in patients with Type 2 diabetes mellitus—a 4-year follow-up study

Aims  Patient education is a very important part of diabetes care. However, until now, little data has been presented about the long‐term effectiveness of structured intensive diabetes education programmes (SIDEP) for people with Type 2 diabetes mellitus.

Cardiovascular autonomic dysfunction predicts acute ischaemic stroke in patients with Type 2 diabetes mellitus : a 7-year follow-up study

Aims  We investigated whether cardiovascular autonomic neuropathy (CAN) is associated with acute ischaemic stroke in patients with Type 2 diabetes.