New Variants Including ARG1 Polymorphisms Associated with C-Reactive Protein Levels Identified by Genome- Wide Association and Pathway Analysis Nadimuthu Vinayagamoorthy 1,2. , Hae-Jin Hu 1,2. , Seon-Hee Yim 1,3 , Seung-Hyun Jung 1,2 , Jaeseong Jo 4 , Sun Ha Jee 4 , Yeun-Jun Chung 1,2 * 1 Integrated Research Center for Genome Polymorphism, The Catholic University of Korea, College of Medicine, Seoul, Korea, 2 Department of Microbiology, The Catholic University of Korea, College of Medicine, Seoul, Korea, 3 Department of Medical Education, The Catholic University of Korea, College of Medicine, Seoul, Korea, 4 Institute of Health Promotion, Graduate School of Public Health, Yonsei University, Seoul, Korea Abstract C-reactive protein (CRP) is a general marker of systemic inflammation and cardiovascular disease (CVD). The genetic contribution to differences in CRP levels remains to be explained, especially in non-European populations. Thus, the aim of this study was to identify genetic loci associated with CRP levels in Korean population. We performed genome-wide association studies (GWAS) using SNPs from 8,529 Korean individuals (7,626 for stage 1 and 903 for stage 2). We also performed pathway analysis. We identified a new genetic locus associated with CRP levels upstream of ARG1 gene (top significant SNP: rs9375813, P meta = 2.85 6 10 28 ), which encodes a key enzyme of the urea cycle counteract the effects of nitric oxide, in addition to known CRP (rs7553007, P meta = 1.72 6 10 216 ) and HNF1A loci (rs2259816, P meta = 2.90 6 10 210 ). When we evaluated the associations between the CRP-related SNPs with cardiovascular disease phenotypes, rs9375813 (ARG1) showed a marginal association with hypertension (P = 0.0440). To identify more variants and pathways, we performed pathway analysis and identified six candidate pathways comprised of genes related to inflammatory processes and CVDs (CRP, HNF1A, PCSK6, CD36, and ABCA1). In addition to the previously reported loci (CRP, HNF1A, and IL6) in diverse ethnic groups, we identified novel variants in the ARG1 locus associated with CRP levels in Korean population and a number of interesting genes related to inflammatory processes and CVD through pathway analysis. Citation: Vinayagamoorthy N, Hu H-J, Yim S-H, Jung S-H, Jo J, et al. (2014) New Variants Including ARG1 Polymorphisms Associated with C-Reactive Protein Levels Identified by Genome-Wide Association and Pathway Analysis. PLoS ONE 9(4): e95866. doi:10.1371/journal.pone.0095866 Editor: Zhi Wei, New Jersey Institute of Technology, United States of America Received September 26, 2013; Accepted March 31, 2014; Published April 24, 2014 Copyright: ß 2014 Vinayagamoorthy et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This study was supported by a grant from the Korea Healthcare Technology R&D Project (A092258), MRC (2012047939), and NRF grant funded by the Korean government (MEST) (2011-0029348), Republic of Korea. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: yejun@catholic.ac.kr . These authors contributed equally to this work. Introduction C-reactive protein (CRP) is an acute phase reactant protein and a general marker of systemic inflammation that is produced by the liver. High CRP levels are known to be associated with cardiovascular disease (CVD) risk factors, including hypertension, coronary heart disease (CHD), and stroke, in addition to traditional risk factors such as BMI, smoking, diabetes, and cholesterol levels [1]. The heritability of CRP levels is estimated to be 25% to 40%, indicating that genetic variations can affect inter-individual or inter-ethnic group differences in CRP levels [2]. Indeed, CRP levels vary significantly among different ethnic groups [3]. For example, serum CRP levels were reported to be relatively lower in East Asians compared to Europeans, South Asians, and Aboriginal peoples in Canada [3]. Several large-scale genome-wide associa- tion studies (GWAS) to identify genetic links to difference in CRP levels have been undertaken; however, most of these studies were performed in European populations [4,5]. In addition to the well- known variants that correlate with CRP levels in Europeans such as CRP, HNF1A (hepatic nuclear factor 1-alpha), and APOE (apolipoprotein E), some recent GWASs have identified new variants such as IL6 (interleukin-6) in the Japanese population and TREM2 (triggering receptors expressed by myeloid cells 2) in African American women [6,7]. Differences in allele frequencies, linkage disequilibrium (LD), effect size, and biological adaptations may influence the identification of variants in different ethnic groups [8]. In spite of the identification of these CRP-associated single- nucleotide polymorphisms (SNPs) and genetic loci by large-scale GWASs, the genetic contributions to differences in CRP levels still need further investigation. Biological pathway-based analyses may be able to obtain more meaningful information from high- throughput whole genome data [8]. Pathway analysis can even suggest candidate variants that might be missed in a classical GWAS approach [9]. In the present study, we used a combined approach of GWAS and pathway analysis and attempted to identify SNPs associated with CRP levels in the Korean population. PLOS ONE | www.plosone.org 1 April 2014 | Volume 9 | Issue 4 | e95866