CCL3L1 Copy Number Research Articles

Copy number variation of CCL3L1 among three major ethnic groups in Malaysia

BackgroundC-C motif Chemokine Ligand 3 Like 1 (CCL3L1) is a multiallelic copy number variable, which plays a crucial role in immunoregulatory and hosts defense through the production of macrophage inflammatory protein (MIP)-1α. Variable range of the CCL3L1 copies from 0 to 14 copies have been documented in several different populations. However, there is still lack of report on the range of CCL3L1 copy number exclusively among Malaysians who are a multi-ethnic population. Thus, this study aims to extensively examine the distribution of CCL3L1 copy number in the three major populations from Malaysia namely Malay, Chinese and Indian. A diploid copy number of CCL3L1 for 393 Malaysians (Malay = 178, Indian = 90, and Chinese = 125) was quantified using Paralogue Ratio Tests (PRTs) and then validated with microsatellites analysis.ResultsTo our knowledge, this is the first report on the CCL3L1 copy number that has been attempted among Malaysians and the Chinese ethnic group exhibits a diverse pattern of CCL3L1 distribution copy number from the Malay and Indian (p < 0.0001). The CCL3L1 ranged from 0 to 8 copies for both the Malay and Indian ethnic groups while 0 to 10 copies for the Chinese ethnic. Consequently, the CCL3L1 copy number among major ethnic groups in the Malaysian population is found to be significantly varied when compared to the European population (p < 0.0001). The mean/median reported for the Malay, Chinese, Indian, and European are 2.759/2.869, 3.453/3.290, 2.437/1.970 and 2.001/1.940 respectively.ConclusionThis study reveals the existence of genetic variation of CCL3L1 in the Malaysian population, and suggests by examining genetic diversity on the ethnicity, and specific geographical region could help in reconstructing human evolutionary history and for the prediction of disease risk related to the CCL3L1 copy number.

Human CCL3L1 copy number variation, gene expression, and the role of the CCL3L1-CCR5 axis in lung function.

Background: The CCL3L1-CCR5 signaling axis is important in a number of inflammatory responses, including macrophage function, and T-cell-dependent immune responses. Small molecule CCR5 antagonists exist, including the approved antiretroviral drug maraviroc,and therapeutic monoclonal antibodies are in development. Repositioning of drugs and targets into new disease areas can accelerate the availability of new therapies and substantially reduce costs. As it has been shown that drug targets with genetic evidence supporting their involvement in the disease are more likely to be successful in clinical development, using genetic association studies to identify new target repurposing opportunities could be fruitful. Here we investigate the potential of perturbation of the CCL3L1-CCR5 axis as treatment for respiratory disease. Europeans typically carry between 0 and 5 copies of CCL3L1 and this multi-allelic variation is not detected by widely used genome-wide single nucleotide polymorphism studies. Methods: We directly measured the complex structural variation of CCL3L1 using the Paralogue Ratio Test and imputed (with validation) CCR5del32 genotypes in 5,000 individuals from UK Biobank, selected from the extremes of the lung function distribution, and analysed DNA and RNAseq data for CCL3L1 from the 1000 Genomes Project. Results: We confirmed the gene dosage effect of CCL3L1 copy number on CCL3L1 mRNA expression levels. We found no evidence for association of CCL3L1 copy number or CCR5del32 genotype with lung function. Conclusions: These results suggest that repositioning CCR5 antagonists is unlikely to be successful for the treatment of airflow obstruction.

Human CCL3L1 copy number variation, gene expression, and the role of the CCL3L1-CCR5 axis in lung function

Background: The CCL3L1-CCR5 signaling axis is important in a number of inflammatory responses, including macrophage function, and T-cell-dependent immune responses. Small molecule CCR5 antagonists exist, including the approved antiretroviral drug maraviroc, and therapeutic monoclonal antibodies are in development. Repositioning of drugs and targets into new disease areas can accelerate the availability of new therapies and substantially reduce costs. As it has been shown that drug targets with genetic evidence supporting their involvement in the disease are more likely to be successful in clinical development, using genetic association studies to identify new target repurposing opportunities could be fruitful. Here we investigate the potential of perturbation of the CCL3L1-CCR5 axis as treatment for respiratory disease. Europeans typically carry between 0 and 5 copies of CCL3L1 and this multi-allelic variation is not detected by widely used genome-wide single nucleotide polymorphism studies. Methods: We directly measured the complex structural variation of CCL3L1 using the Paralogue Ratio Test and imputed (with validation) CCR5d32 genotypes in 5,000 individuals from UK Biobank, selected from the extremes of the lung function distribution, and analysed DNA and RNAseq data for CCL3L1 from the 1000 Genomes Project. Results: We confirmed the gene dosage effect of CCL3L1 copy number on CCL3L1 mRNA expression levels. We found no evidence for association of CCL3L1 copy number or CCR5d32 genotype with lung function. Conclusions: These results suggest that repositioning CCR5 antagonists is unlikely to be successful for the treatment of airflow obstruction.

Human CCL3L1 copy number variation, gene expression, and the role of the CCL3L1-CCR5 axis in lung function

Background: The CCL3L1-CCR5 signaling axis is important in a number of inflammatory responses, including macrophage function, and T-cell-dependent immune responses. Small molecule CCR5 antagonists exist, including the approved antiretroviral drug maraviroc, and therapeutic monoclonal antibodies are in development. Repositioning of drugs and targets into new disease areas can accelerate the availability of new therapies and substantially reduce costs. As it has been shown that drug targets with genetic evidence supporting their involvement in the disease are more likely to be successful in clinical development, using genetic association studies to identify new target repurposing opportunities could be fruitful. Here we investigate the potential of perturbation of the CCL3L1-CCR5 axis as treatment for respiratory disease. Europeans typically carry between 0 and 5 copies of CCL3L1 and this multi-allelic variation is not detected by widely used genome-wide single nucleotide polymorphism studies. Methods: We directly measured the complex structural variation of CCL3L1 using the Paralogue Ratio Test and imputed (with validation) CCR5del32 genotypes in 5,000 individuals from UK Biobank, selected from the extremes of the lung function distribution, and analysed DNA and RNAseq data for CCL3L1 from the 1000 Genomes Project. Results: We confirmed the gene dosage effect of CCL3L1 copy number on CCL3L1 mRNA expression levels. We found no evidence for association of CCL3L1 copy number or CCR5del32 genotype with lung function. Conclusions: These results suggest that repositioning CCR5 antagonists is unlikely to be successful for the treatment of airflow obstruction.

CD41 T cell recovery during suppression of HIV replication: an international comparison of the immunological efficacy of antiretroviral therapy in North America, Asia and Africa.

Even among HIV-infected patients who fully suppress plasma HIV RNA replication on antiretroviral therapy, genetic (e.g. CCL3L1 copy number), viral (e.g. tropism) and environmental (e.g. chronic exposure to microbial antigens) factors influence CD4 recovery. These factors differ markedly around the world and therefore the expected CD4 recovery during HIV RNA suppression may differ globally. We evaluated HIV-infected adults from North America, West Africa, East Africa, Southern Africa and Asia starting non-nucleoside reverse transcriptase inhibitorbased regimens containing efavirenz or nevirapine, who achieved at least one HIV RNA level <500/ml in the first year of therapy and observed CD4 changes during HIV RNA suppression. We used a piecewise linear regression to estimate the influence of region of residence on CD4 recovery, adjusting for socio-demographic and clinical characteristics. We observed 28 217 patients from 105 cohorts over 37 825 person-years. After adjustment, patients from East Africa showed diminished CD4 recovery as compared with other regions. Three years after antiretroviral therapy initiation, the mean CD4 count for a prototypical patient with a pre-therapy CD4 count of 150/ml was 529/ml [95% confidence interval (CI): 517–541] in North America, 494/ml (95% CI: 429–559) in West Africa, 515/ml (95% CI: 508–522) in Southern Africa, 503/ml (95% CI: 478–528) in Asia and 437/ml (95% CI: 425–449) in East Africa. CD4 recovery during HIV RNA suppression is diminished in East Africa as compared with other regions of the world, and observed differences are large enough to potentially influence clinical outcomes. Epidemiological analyses on a global scale can identify macroscopic effects unobservable at the clinical, national or individual regional level.

Gene copy number variation in Indian population and its implication in health

Objectives Copy number variations (CNV) are important source of human genetic variation, found to be widely prevalent than what was initially predicted. The involvement of CNVs in disease susceptibility and drug response is well reported in other populations but has not been studied to a larger extent in Indian population. The aim of the present study was to evaluate the distribution of copy number variable genes in Indian population and their link, if any, to health, disease and drug response along with development of a comprehensive resource of CNV frequency distribution among different populations. Methods A total of 100 – 280 healthy controls and in variable number of patients from Indian population were genotyped. Genotyping of the copy number (CN) variable genes was carried by PCR-based methodologies (long range PCR, real-time PCR and PRT assay). Results An indicative correlation with disease susceptibility and significant (p < 0.05) difference in the frequency distribution of the CNV variants was observed in our study. MTUS1 deletion variant was found to be significantly (p = 0.0207) associated with a decreased risk to breast cancer. A significant association between CCL3L1 copy number and risk to HIV-1 was also observed. The interethnic comparison of CCL3L1 gene copy number frequency demonstrated significant difference worldwide, being highest in Africans. The common deletion frequency comprising LCE3B and LCE3C genes was found to be highest in European and American populations. The frequency of FCGR3B CN < 2 was found comparatively higher in Indians, Americans and Africans as compared to European Caucasians. Thus, inter-ethnic differences were observed among populations, highlighting varied frequency distribution pattern based on their distinct geographical location. Conclusion This is our first attempt to create a comprehensive map of the frequency distribution of CNVs and its association to disease susceptibility in Indian population. This varied worldwide distribution can thus be relevant from clinical or evolutionary point of view in future.

CCL3L1 copy number, HIV load, and immune reconstitution in sub-Saharan Africans

BackgroundThe role of copy number variation of the CCL3L1 gene, encoding MIP1α, in contributing to the host variation in susceptibility and response to HIV infection is controversial. Here we analyse a sub-Saharan African cohort from Tanzania and Ethiopia, two countries with a high prevalence of HIV-1 and a high co-morbidity of HIV with tuberculosis.MethodsWe use a form of quantitative PCR called the paralogue ratio test to determine CCL3L1 gene copy number in 1134 individuals and validate our copy number typing using array comparative genomic hybridisation and fiber-FISH.ResultsWe find no significant association of CCL3L1 gene copy number with HIV load in antiretroviral-naïve patients prior to initiation of combination highly active anti-retroviral therapy. However, we find a significant association of low CCL3L1 gene copy number with improved immune reconstitution following initiation of highly active anti-retroviral therapy (p = 0.012), replicating a previous study.ConclusionsOur work supports a role for CCL3L1 copy number in immune reconstitution following antiretroviral therapy in HIV, and suggests that the MIP1α -CCR5 axis might be targeted to aid immune reconstitution.

CCR5 Haplotypes Influence HCV Serostatus in Caucasian Intravenous Drug Users

BackgroundUp to 90% HIV-1 positive intravenous drug users (IDUs) are co-infected with HCV. Although best recognized for its function as a major co-receptor for cell entry of HIV, CC chemokine receptor 5 (CCR5) has also been implicated in the pathogenesis of HCV infection. Here, we investigated whether CCR5 haplotypes influence HIV-1 and HCV seropositivity among 373 Caucasian IDUs from Estonia.MethodsOf these IDUs, 56% and 44% were HIV and HCV seropositive, respectively, and 47% were coinfected. 500 blood donors seronegative for HIV and HCV were also evaluated. CCR5 haplotypes (HHA to HHG*2) were derived after genotyping nine CCR2–CCR5 polymorphisms. The association between CCR5 haplotypes with HIV and/or HCV seropositivity was determined using logistic regression analysis. Co-variates included in the models were length of intravenous drug use, HBV serostatus and copy number of CCL3L1, the gene encoding the most potent HIV-suppressive chemokine and ligand for CCR5.ResultsCompared to IDUs seronegative for both HCV and HIV (HCV−/HIV-), IDUs who were HCV+/HIV- and HCV+/HIV+were 92% and 82%, respectively, less likely to possess the CCR5-HHG*1 haplotype, after controlling for co-variates (Padjusted = 1.89×10−4 and 0.003, respectively). This association was mostly due to subjects bearing the CCR5 HHE and HHG*1 haplotype pairs. Approximately 25% and<10% of HCV−/HIV- IDUs and HCV−/HIV- blood donors, respectively, possessed the HHE/HHG*1 genotype.ConclusionsOur findings suggest that HHG*1-bearing CCR5 genotypes influence HCV seropositivity in a group of Caucasian IDUs.

Functional effects of CCL3L1 copy number

Copy number variation (CNV) is becoming increasingly important as a feature of human variation in disease susceptibility studies. However, the consequences of copy number variation are not so well understood. Here we present data exploring the functional consequences of copy number variation of CCL3L1 in 55 independent UK samples with no known clinical phenotypes. Copy number of CCL3L1 was determined by the paralogue ratio test (PRT), and expression levels of MIP-1α and mRNA from stimulated monocytes were measured and analysed. The data show no statistically significant association of MIP-1α protein levels with copy number. However, there was a significant correlation between copy number and CCL3L1:CCL3 mRNA ratio. The data also provide evidence that expression of CCL3 predominates in both protein and mRNA, and therefore the observed variation of CCL3 is potentially more important biologically than that of copy number variation of CCL3L1.

CCL3L1 copy number and susceptibility to malaria

Copy number variation can contribute to the variation observed in susceptibility to complex diseases. Here we present the first study to investigate copy number variation of the chemokine gene CCL3L1 with susceptibility to malaria. We present a family-based genetic analysis of a Tanzanian population (n=922), using parasite load, mean number of clinical infections of malaria and haemoglobin levels as phenotypes. Copy number of CCL3L1 was measured using the paralogue ratio test (PRT) and the dataset exhibited copy numbers ranging between 1 and 10 copies per diploid genome (pdg). Association between copy number and phenotypes was assessed. Furthermore, we were able to identify copy number haplotypes in some families, using microsatellites within the copy variable region, for transmission disequilibrium testing. We identified a high level of copy number haplotype diversity and find some evidence for an association of low CCL3L1 copy number with protection from anaemia.

509 CCL3L1 Protein Did Not Affect IL-6 Expression, but Significantly Up-regulated IL-10 Expression in the Allergic Response

BackgroundPreviously, we found that the mean copy number of CCL3L1 in patients with asthma was significantly lower than that of control subjects (3.13 vs 3.75, P = 0.001). We investigated its possible molecular mechanism using a human monocytic cell line stimulated with house dust mite extract.MethodThe THP-1 human monocytic cells were stimulated with various concentrations of HDM extract. After stimulation, assay-on-demand gene expression products (Applied Biosystems) were used to evaluate mRNA expression of CCL3L1 (Hs 00609691_ml), IlL-6 (Hs00174131_ml), and IL-10 (Hs00961622_ml) levels as measurement of mRNA levels by real time PCR.ResultsTreatment of THP-1 cells with various concentration of HDM extract induced marked up- regulation of the expression of cytokines IL-10 and IL-6, which indicated that allergic responses were efficiently induced. Recombinant CCL3L1 protein had no effect on cytokine expression of THP-1 Cells in absence of HDM extract stimulation. In the presence of HDM extract (10 ug/mL) stimulation, CCL3L1 protein significantly up-regulated IL-10 expression (Ratio to ng/mL CCL3L1) dose-dependantly (0 ug/mL CCL3L1 + 0.3;12.4, 10 ug/mL CCL3L1; 15.8+ 1.1, 50 ug/mL CCL3L1; 16.8+ 0.3, 100 ug/mL CCL3L1; 18.0+ 0.8, (P > 0.05), but did not affect IL-6 expression (P > 0.05).ConclusionThe significantly elevated asthma risk in subjects with a low copy number of the CCL3L1 gene which may be down-regulating IL-10 expression, not IL-6 expression.

CCL3L1 gene copy number in individuals with and without HIV-associated neurocognitive disorder

CCL3L1 copy number variation has been implicated as a marker for susceptibility and immunity to human immunodeficiency virus (HIV)-1 infection and its pathogenic sequelae. Some of these findings have been confirmed in several, but not all, subsequent independent cohort studies. A three-fold risk for the development of HIV-associated dementia was reported in individuals possessing a CCL3L1 copy number below the ethnic group median combined with a detrimental CCR5 genotype. With the availability of antiretroviral therapy since 1996, there has been a significant decline in HIV-associated dementia, and milder forms of HIV-associated neurocognitive impairment (HAND) are now most prevalent. Moreover, patients are living longer with HIV-1 infection and it is recognized that aging may be a contributory factor to the development of cognitive disorder. Thus, the need for biomarkers that can be used in clinical practice to identify and provide optimal treatment for those at increased risk for HAND is great. HAND affects 20%-30% of HIV-infected individuals, and several genetic loci which have been shown to confer susceptibility to HIV infection may also modulate the development of neurocognitive disorder. The aim of this study was to determine whether CCL3L1 chemokine gene copy number in self-defined ethnic groups could differentiate HIV-infected individuals with and without HAND. Genomic DNA was isolated from buccal swabs or peripheral blood mononuclear cells obtained from HIV-infected patients with or without a diagnoses of neurocognitive dysfunction in the Northeast AIDS Dementia Cohort and National NeuroAIDS Tissue Consortium. To maintain a uniform standard, a quantitative polymerase chain reaction design similar to previous studies using Taqman probes and fixed input DNA between 2 ng and 10 ng was used to determine a CCL3L1 copy number. Standard curves with two-fold dilutions from 25 ng to 1.56 ng were generated. CCL3L1 copy number was determined in triplicate in 262 subjects using quantitative polymerase chain reaction and the relative quantitation method. Data were analyzed using analysis of variance, with significance defined as P < 0.05 and Bonferroni post hoc tests. Significant differences as determined by analysis of variance in CCL3L1 copy number between African-Americans and Caucasians (P < 0.0001) were found, highlighting ethnic group differences in the copy number of this gene. However, there were no differences in CCL3L1 copy number across the neurocognitive groups within each ethnic group. The median CCL3L1 copy number in African-Americans of two and Caucasians of one in this study was significantly lower than the previously reported ethnic group means of two and four copies, respectively. A higher prevalence of abnormal cognition with a relative risk of four was seen in African-Americans versus Caucasians. Based on this nested case-control study, CCL3L1 copy number alone may not be useful for distinguishing between individuals at risk for mild or severe neurocognitive disorder. Additional larger cohort studies are required to determine whether CCL3L1 copy number in combination with polymorphisms in other genes known to contribute to HIV risk will be useful in identifying those at increased risk for HAND.

Copy number variation in CCL3L1 gene is associated with susceptibility to acute rejection in patients after liver transplantation

Copy number variation (CNV) in gene was a novel type of genetic diversity in human that affect the pathogenesis, progress, and prognosis of disease. The aim of this study was to determine the associations between CNV in CCL3L1 gene and the rejection risk in liver-transplant recipients. The 266 Han-Chinese patients and 135 volunteers were enrolled in this study. Genomic DNA and mRNA samples were obtained from the whole peripheral blood; a quantitative real-time PCR-based assay was carried out to determine the copy number of CCL3L1, and real-time PCR was carried out to calculate the CCL3L1/CCL3 mRNA ratio. The CNV distributions in patients and health controls had no significant differences. Copy number in acute rejection group (AR) shifted to higher number compared with non-acute rejection group (4.74±1.87 vs. 3.88±2.13, p=0.017). Moreover, patients with higher CCL3L1 copies (≥3 copies) had a significantly higher rejection risk than patients lower copies CCL3L1 (OR=3.85, 95% CI, 1.14-13.04; p=0.021). No association was found between CNV and rejection grades. In conclusion, liver transplant recipients with high copy number of CCL3L1 gene had a significant higher risk of AR. CNV might be a novel genetic diversity that correlated with allograft rejection.

Accuracy and differential bias in copy number measurement of CCL3L1 in association studies with three auto-immune disorders

BackgroundCopy number variation (CNV) contributes to the variation observed between individuals and can influence human disease progression, but the accurate measurement of individual copy numbers is technically challenging. In the work presented here we describe a modification to a previously described paralogue ratio test (PRT) method for genotyping the CCL3L1/CCL4L1 copy variable region, which we use to ascertain CCL3L1/CCL4L1 copy number in 1581 European samples. As the products of CCL3L1 and CCL4L1 potentially play a role in autoimmunity we performed case control association studies with Crohn's disease, rheumatoid arthritis and psoriasis clinical cohorts.ResultsWe evaluate the PRT methodology used, paying particular attention to accuracy and precision, and highlight the problems of differential bias in copy number measurements. Our PRT methods for measuring copy number were of sufficient precision to detect very slight but systematic differential bias between results from case and control DNA samples in one study. We find no evidence for an association between CCL3L1 copy number and Crohn's disease, rheumatoid arthritis or psoriasis.ConclusionsDifferential bias of this small magnitude, but applied systematically across large numbers of samples, would create a serious risk of false positive associations in copy number, if measured using methods of lower precision, or methods relying on single uncorroborated measurements. In this study the small differential bias detected by PRT in one sample set was resolved by a simple pre-treatment by restriction enzyme digestion.

Copy number variation of CCL3L1 influences asthma risk by modulating IL-10 expression

BackgroundCopy number of Chemokine ligand 3-like 1 (CCL3L1) is associated with various immune disorders. This study was conducted to assess the role of CCL3L1 in asthma by both association analyses of human subjects and in vitro functional analyses. MethodsWe analyzed the copy number of the CCL3L1 gene in 533 Korean subjects (372 controls and 161 asthma patients) by real-time PCR, and investigated the effect of recombinant CCL3L1 protein on THP-1 human monocytic cells that were stimulated with house dust mite extract. ResultsThe mean copy number of CCL3L1 in asthma subjects was significantly lower than that of control subjects (3.18 vs. 3.75, p=0.001). A low copy number of ≤1 was significantly associated with increased asthma risk with an odds ratio of 2.47, and a high copy number of ≥5 was associated with decreased asthma risk with an odds ratio of 0.40. Subjects with ≤1 copy of CCL3L1 had significantly lower mRNA levels of CCL3L1 in peripheral blood cells, and significantly higher serum IgE levels (p<0.05). In the house dust mite extract-simulated THP-1 monocytic cells, CCL3L1 protein dose-dependently up-regulated the expression of IL-10, an anti-inflammatory cytokine. ConclusionCopy number of CCL3L1 may influence asthma risk by modulating IL-10 expression.

Influence of Variations in CCL3L1 and CCR5 on Tuberculosis in a Northwestern Colombian Population

We investigated the association of polymorphisms in CCR5, the major human immunodeficiency virus (HIV)-1 coreceptor, and copy number of its potent ligand CCL3L1 with tuberculosis in 298 individuals from Colombia. The CCR5-HHD haplotype, a known genetic determinant of increased susceptibility to HIV-AIDS, and a high copy number of CCL3L1, a known genetic determinant of enhanced CCL3/CCL3L1 chemokine expression, each associated with presence of tuberculosis. Furthermore, CCR5-HHD was associated with higher CCR5 gene and surface expression. These results substantiate the strong link between the pro-inflammatory effects of CCR5 and its ligands with active tuberculosis and suggest that chemokine-chemokine receptor genetic determinants may influence tuberculosis in addition to HIV/AIDS.

An Additive Effect of Protective Host Genetic Factors Correlates With HIV Nonprogression Status

In HIV-positive individuals, complex multifactorial mechanisms control viral infection. In addition to viral and immunological factors, the host genetic background also plays an important role. Our objective was to evaluate how various genetic factors associated with delayed AIDS onset. Thirty HIV+ long-term nonprogressors (LTNPs) and 30 known progressors were analyzed. Host genes were analyzed in peripheral blood mononuclear cells DNA: CCR5 and HLA were polymerase chain reaction typed. HLA-C5', HCP5 polymorphisms, and CCL3L1 copy number were determined using real-time polymerase chain reaction. The CCL3L1high-copy-CCR5 deletion genetic risk groups was overrepresented in LTNPs. However, separately, neither CCL3L1 nor CCR5 were significantly associated with clinical outcome. HLA seemed as a strong nonprogression determinant, mainly HLA-B and the less-studied HLA-C. HLA-Cw0102 and HLA-C5' had an impact on LTNP phenotype along with HLA-B5701 and B2705. The presence of allele combinations like HLA- B*5701-Cw0602, HLA-B*2705-Cw0102, or HLA-B*3801-Cw1203 had the strongest effect in non-progression. As for HCP5, no independent effect was observed. The studied factors had additive effects, and although the number of patients was small, it seemed that carrying a high number of protective alleles associated with progression delay. We showed the additive load of protective host factors was predictive of nonprogression, and that HLA-associated factors were predominant in this global effect.

CCL3L1 copy number variation and susceptibility to HIV-1 infection: a meta-analysis.

BackgroundAlthough several studies have investigated whether CCL3L1 copy number variation (CNV) influences the risk of HIV-1 infection, there are still no clear conclusions. Therefore, we performed a meta-analysis using two models to generate a more robust estimate of the association between CCL3L1 CNV and susceptibility to HIV-1 infection.MethodsWe divided the cases and controls into two parts as individuals with CCL3L1 gene copy number (GCN) above the population specific median copy number (PMN) and individuals with CCL3L1 GCN below PMN, respectively. Odds ratios (ORs) with 95% confidence intervals (95% CIs) were given for the main analysis. We also conducted stratified analyses by ethnicity, age group and sample size. Relevant literatures were searched through PubMed and ISI Web of Knowledge up to March 2010.ResultsIn total, 9 studies with 2434 cases and 4029 controls were included. ORs for the main analysis were 1.35 (95% CI, 1.02–1.78, model: GCN ≤ PMN Vs. GCN > PMN) and 1.70 (95% CI, 1.30–2.23, model: GCN < PMN Vs. GCN ≥ PMN), respectively. Either in stratified analysis, statistically significant results can be detected in some subgroups.ConclusionsOur analyses indicate that CCL3L1 CNV is associated with susceptibility to HIV-1 infection. A lower copy number is associated with an increased risk of HIV-1 infection, while a higher copy number is associated with reduced risk for acquiring HIV-1.

Evolution of haplotypes at CCL3L1/CCL4L1

Background CCL3L1 and CCL4L1 are chemokine genes, located on chromosome 17q12 (Figure 1). They are copy number variable genes that share 95% sequence identity with their non-copy number variable paralogues CCL3 and CCL4 [1]. The copy number (CN) of these genes varies between populations [2] and has been shown to be associated with phenotypes, such as susceptibility to HIV infection [2] and SLE [3]. A CCL3L1 pseudogene, also known as CCL3L2, is present in the CCL3L1 region. This pseudogene has sequences similar to CCL3L1 gene, but lacks exon 1 of CCL3L1. As a result, its presence might affect copy number (CN) measurement and subsequent interpretations in association studies between CCL3L1 CN and diseases [4]. The copy number of CCL3L1/CCL4L1 was measured using paralogue ratio test (PRT) in 270 HapMap samples 192 UK samples and 157 Basques samples [5]. Firstly, we examined the association between the presence of the CCL3L1 pseudogene and CCL3L1 CN in the UK samples and HapMap samples by PCR. The pseudogene was found in 52 out of 192 (27.08%) of UK samples. The presence of this pseudogene is strongly associated with higher copy number of CCL3L1/CCL4L1 (P<1 × 10) (Figure 2). The presence of the pseudogene was tested in all HapMap populations (Table 1). SNP genotyping and CCL3 microsatellite assays were then carried out to define a set of flanking markers that may predict CCL3L1/ CCL4L1 CN in UK and Basque samples. The best combination of 2 flanking SNPs, rs16972085 and rs8064426, can be used to predict the CCL3L1/CCL4L1 CN in UK and Basque samples with only 70% accuracy. Although the CCL3 microsatellite alleles are not associated with CCL3L1/CCL4L1 copy number, there is extensive allelic diversity in the microsatellite. Finally, to improve the accuracy of CCL3L1/CCL4L1 CN prediction, the CCL3L1/CCL4L1 genes were sequenced in 90 CEU samples to identify sequence variants within the copy-variable genes themselves. Analysis of CCL3L1/CCL41 haplotypes in CEU samples is underway to provide information on evolution of the CCL3L1/CCL4L1 haplotypes and the relationship between these haplotypes, flanking SNPs and the presence of the CCL3L1 pseudogene.

Association of CCR2-CCR5 Haplotypes and CCL3L1 Copy Number with Kawasaki Disease, Coronary Artery Lesions, and IVIG Responses in Japanese Children

BackgroundThe etiology of Kawasaki Disease (KD) is enigmatic, although an infectious cause is suspected. Polymorphisms in CC chemokine receptor 5 (CCR5) and/or its potent ligand CCL3L1 influence KD susceptibility in US, European and Korean populations. However, the influence of these variations on KD susceptibility, coronary artery lesions (CAL) and response to intravenous immunoglobulin (IVIG) in Japanese children, who have the highest incidence of KD, is unknown.Methodology/Principal FindingsWe used unconditional logistic regression analyses to determine the associations of the copy number of the CCL3L1 gene-containing duplication and CCR2-CCR5 haplotypes in 133 Japanese KD cases [33 with CAL and 25 with resistance to IVIG] and 312 Japanese controls without a history of KD. We observed that the deviation from the population average of four CCL3L1 copies (i.e., < or > four copies) was associated with an increased risk of KD and IVIG resistance (adjusted odds ratio (OR) = 2.25, p = 0.004 and OR = 6.26, p = 0.089, respectively). Heterozygosity for the CCR5 HHF*2 haplotype was associated with a reduced risk of both IVIG resistance (OR = 0.21, p = 0.026) and CAL development (OR = 0.44, p = 0.071).Conclusions/SignificanceThe CCL3L1-CCR5 axis may play an important role in KD pathogenesis. In addition to clinical and laboratory parameters, genetic markers may also predict risk of CAL and resistance to IVIG.