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Leu432Val polymorphism in CYP1B1 as a susceptible factor towards predisposition to primary open-angle glaucoma.

Bhattacharjee A, Banerjee D, Mookherjee S, Acharya M, Banerjee A, Ray A, Sen A, Indian Genome Variation ConsortiumRay K - Mol. Vis. (2008)

Bottom Line: CYP1B1 Val432 was estimated to generate higher ROS in RPE cells compared to its allelic variant (Leu432; p=0.0245 for 15 min and p=0.0197 for 30 min).Comparison of haplotype diversities revealed CGGTA as the risk haplotype for the disease (p=0.0001, by Fisher's exact test).Higher ROS generation by Val432 in CYP1B1 might lead to apoptotic change that leads to glaucoma.

View Article: PubMed Central - PubMed

Affiliation: Molecular and Human Genetics Division, Indian Institute of Chemical Biology (Council of Scientific & Industrial Research), Kolkata, India.

ABSTRACT

Purpose: Defects in cytochrome P450 1B1 (CYP1B1) cause primary congenital glaucoma. However, defects in the gene have also been reported in primary open-angle glaucoma (POAG). Since POAG is primarily a complex disease, we examined the potential of coding single nucleotide polymorphisms (cSNPs) in the gene for association with the disease.

Methods: Five coding SNPs - c.514 C>G (Arg48Gly), c.727 G>T (Ala119Ser), c.1666 C>G (Leu432Val), c.1719 C>T (Asp449Asp), and c.1730 A>G (Asn453Ser) - were genotyped in 264 unrelated POAG patients and 95 controls. In addition, 542 normal individuals selected from various ethnic groups representing the Indian population were also genotyped for these cSNPs. The patterns of linkage disequilibrium between the SNPs and haplotype variations for comparison between POAG patients and controls as well as different ethnic groups of the Indian population were determined using Haploview. Allelic variants of Leu432Val were cloned by site-directed mutagenesis of normal CYP1B1 cDNA, which were used for transfection of retinal pigment epithelium (RPE) cells. The generation of reactive oxygen species (ROS) was quantified by measuring fluorescence emission by degradation of CM-H2DCFDA using a fluoremeter.

Results: The c.1666G allele of the Leu432Val in CYP1B1 showed a statistically significant higher representation among POAG patients compared to controls (p=0.0001; Odds ratio=6.027; 95% CI: 3.863-9.401) suggesting it to be a potential risk allele toward disease predisposition. Analysis of genotype frequencies of the polymorphism between the two groups demonstrated GG as a potential risk genotype (p=0.0001; Odds ratio=15.505; 95% CI: 5.529-43.474) for the disease. CYP1B1 Val432 was estimated to generate higher ROS in RPE cells compared to its allelic variant (Leu432; p=0.0245 for 15 min and p=0.0197 for 30 min). Comparison of haplotype diversities revealed CGGTA as the risk haplotype for the disease (p=0.0001, by Fisher's exact test).

Conclusions: We report CYP1B1 c.1666G (Val432) as a susceptible allele for POAG and CGGTA as the risk haplotype for the disease. Higher ROS generation by Val432 in CYP1B1 might lead to apoptotic change that leads to glaucoma. Remarkable variation of the cSNPs observed among ethnic groups of India could provide insight for future epidemiological studies on POAG in these population groups.

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CYP1B1 haplotype distribution. A: CYP1B1 haplotype variation among primary open-angle glaucoma patients and controls The Study of haplotype diversity among patients and controls revealed C-G-G-T-A as the predominant haplotype among POAG patients (41%) and a lower representation among controls (10%). Hence, the haplotype represents a potential risk haplotype among POAG patients. B: The distribution of the risk haplotype (CGGTA) in 24 ethnic groups of the Indian population. The study on the distribution of the risk haplotype (C-G-G-T-A) in the various ethnic groups of the Indian population revealed the highest frequency (26.6%) among the ethnic group, IE-N-LP5, followed by AA-C-IP5 (25%). In the OG-W-LP5 group where c.1666G (Val432) is the major allele, the risk haplotype frequency was estimated to be 16.7%.
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f5: CYP1B1 haplotype distribution. A: CYP1B1 haplotype variation among primary open-angle glaucoma patients and controls The Study of haplotype diversity among patients and controls revealed C-G-G-T-A as the predominant haplotype among POAG patients (41%) and a lower representation among controls (10%). Hence, the haplotype represents a potential risk haplotype among POAG patients. B: The distribution of the risk haplotype (CGGTA) in 24 ethnic groups of the Indian population. The study on the distribution of the risk haplotype (C-G-G-T-A) in the various ethnic groups of the Indian population revealed the highest frequency (26.6%) among the ethnic group, IE-N-LP5, followed by AA-C-IP5 (25%). In the OG-W-LP5 group where c.1666G (Val432) is the major allele, the risk haplotype frequency was estimated to be 16.7%.

Mentions: Haplotypes were constructed using Haploview with the studied cSNPs in the following order: c.514 C>G, c.727 G>T, c.1666 C>G, c.1719 C>T, and c.1730 A>G. Study of haplotype diversity (Figure 5) among patients and controls revealed C-G-G-T-A as the predominant haplotype among POAG patients (41%) and a lower representation among controls (10%; p=0.0001, by Fisher’s exact test; Table 3). Hence, the haplotype represents a potential risk haplotype among POAG patients. Interestingly, this haplotype has been reported to harbor a vast majority of the CYP1B1 mutations causal to primary congenital glaucoma (PCG) across different population groups worldwide [35]. Therefore, the risk haplotype for POAG as revealed by our study also represents the risk haplotype for PCG. However, among the nine POAG patients harboring CYP1B1 mutations [20], we could determine haplotype in only two patients with access to family members’ samples, but the risk haplotype (C-G-G-T-A) was not identified in either of them. Incidentally, we observed that the haplotype of the wild type CYP1B1 cDNA clone (c.1666 C variant), used to examine functional variation of Leu432Val polymorphism, is the same as the most common haplotype (C-G-C-C-A) present in 40% of the controls examined (Table 3). The risk haplotype (C-G-G-T-A), which is overrepresented in the POAG patients, is functionally equivalent to the haplotype (C-G-G-C-A), which is present in the mutant clone, since the cSNP (c.1719 C>T) in LD with Leu432Val (c.1666 C>G) polymorphism codes for a synonymous change (Asp449Asp).


Leu432Val polymorphism in CYP1B1 as a susceptible factor towards predisposition to primary open-angle glaucoma.

Bhattacharjee A, Banerjee D, Mookherjee S, Acharya M, Banerjee A, Ray A, Sen A, Indian Genome Variation ConsortiumRay K - Mol. Vis. (2008)

CYP1B1 haplotype distribution. A: CYP1B1 haplotype variation among primary open-angle glaucoma patients and controls The Study of haplotype diversity among patients and controls revealed C-G-G-T-A as the predominant haplotype among POAG patients (41%) and a lower representation among controls (10%). Hence, the haplotype represents a potential risk haplotype among POAG patients. B: The distribution of the risk haplotype (CGGTA) in 24 ethnic groups of the Indian population. The study on the distribution of the risk haplotype (C-G-G-T-A) in the various ethnic groups of the Indian population revealed the highest frequency (26.6%) among the ethnic group, IE-N-LP5, followed by AA-C-IP5 (25%). In the OG-W-LP5 group where c.1666G (Val432) is the major allele, the risk haplotype frequency was estimated to be 16.7%.
© Copyright Policy - open-access
Related In: Results  -  Collection

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f5: CYP1B1 haplotype distribution. A: CYP1B1 haplotype variation among primary open-angle glaucoma patients and controls The Study of haplotype diversity among patients and controls revealed C-G-G-T-A as the predominant haplotype among POAG patients (41%) and a lower representation among controls (10%). Hence, the haplotype represents a potential risk haplotype among POAG patients. B: The distribution of the risk haplotype (CGGTA) in 24 ethnic groups of the Indian population. The study on the distribution of the risk haplotype (C-G-G-T-A) in the various ethnic groups of the Indian population revealed the highest frequency (26.6%) among the ethnic group, IE-N-LP5, followed by AA-C-IP5 (25%). In the OG-W-LP5 group where c.1666G (Val432) is the major allele, the risk haplotype frequency was estimated to be 16.7%.
Mentions: Haplotypes were constructed using Haploview with the studied cSNPs in the following order: c.514 C>G, c.727 G>T, c.1666 C>G, c.1719 C>T, and c.1730 A>G. Study of haplotype diversity (Figure 5) among patients and controls revealed C-G-G-T-A as the predominant haplotype among POAG patients (41%) and a lower representation among controls (10%; p=0.0001, by Fisher’s exact test; Table 3). Hence, the haplotype represents a potential risk haplotype among POAG patients. Interestingly, this haplotype has been reported to harbor a vast majority of the CYP1B1 mutations causal to primary congenital glaucoma (PCG) across different population groups worldwide [35]. Therefore, the risk haplotype for POAG as revealed by our study also represents the risk haplotype for PCG. However, among the nine POAG patients harboring CYP1B1 mutations [20], we could determine haplotype in only two patients with access to family members’ samples, but the risk haplotype (C-G-G-T-A) was not identified in either of them. Incidentally, we observed that the haplotype of the wild type CYP1B1 cDNA clone (c.1666 C variant), used to examine functional variation of Leu432Val polymorphism, is the same as the most common haplotype (C-G-C-C-A) present in 40% of the controls examined (Table 3). The risk haplotype (C-G-G-T-A), which is overrepresented in the POAG patients, is functionally equivalent to the haplotype (C-G-G-C-A), which is present in the mutant clone, since the cSNP (c.1719 C>T) in LD with Leu432Val (c.1666 C>G) polymorphism codes for a synonymous change (Asp449Asp).

Bottom Line: CYP1B1 Val432 was estimated to generate higher ROS in RPE cells compared to its allelic variant (Leu432; p=0.0245 for 15 min and p=0.0197 for 30 min).Comparison of haplotype diversities revealed CGGTA as the risk haplotype for the disease (p=0.0001, by Fisher's exact test).Higher ROS generation by Val432 in CYP1B1 might lead to apoptotic change that leads to glaucoma.

View Article: PubMed Central - PubMed

Affiliation: Molecular and Human Genetics Division, Indian Institute of Chemical Biology (Council of Scientific & Industrial Research), Kolkata, India.

ABSTRACT

Purpose: Defects in cytochrome P450 1B1 (CYP1B1) cause primary congenital glaucoma. However, defects in the gene have also been reported in primary open-angle glaucoma (POAG). Since POAG is primarily a complex disease, we examined the potential of coding single nucleotide polymorphisms (cSNPs) in the gene for association with the disease.

Methods: Five coding SNPs - c.514 C>G (Arg48Gly), c.727 G>T (Ala119Ser), c.1666 C>G (Leu432Val), c.1719 C>T (Asp449Asp), and c.1730 A>G (Asn453Ser) - were genotyped in 264 unrelated POAG patients and 95 controls. In addition, 542 normal individuals selected from various ethnic groups representing the Indian population were also genotyped for these cSNPs. The patterns of linkage disequilibrium between the SNPs and haplotype variations for comparison between POAG patients and controls as well as different ethnic groups of the Indian population were determined using Haploview. Allelic variants of Leu432Val were cloned by site-directed mutagenesis of normal CYP1B1 cDNA, which were used for transfection of retinal pigment epithelium (RPE) cells. The generation of reactive oxygen species (ROS) was quantified by measuring fluorescence emission by degradation of CM-H2DCFDA using a fluoremeter.

Results: The c.1666G allele of the Leu432Val in CYP1B1 showed a statistically significant higher representation among POAG patients compared to controls (p=0.0001; Odds ratio=6.027; 95% CI: 3.863-9.401) suggesting it to be a potential risk allele toward disease predisposition. Analysis of genotype frequencies of the polymorphism between the two groups demonstrated GG as a potential risk genotype (p=0.0001; Odds ratio=15.505; 95% CI: 5.529-43.474) for the disease. CYP1B1 Val432 was estimated to generate higher ROS in RPE cells compared to its allelic variant (Leu432; p=0.0245 for 15 min and p=0.0197 for 30 min). Comparison of haplotype diversities revealed CGGTA as the risk haplotype for the disease (p=0.0001, by Fisher's exact test).

Conclusions: We report CYP1B1 c.1666G (Val432) as a susceptible allele for POAG and CGGTA as the risk haplotype for the disease. Higher ROS generation by Val432 in CYP1B1 might lead to apoptotic change that leads to glaucoma. Remarkable variation of the cSNPs observed among ethnic groups of India could provide insight for future epidemiological studies on POAG in these population groups.

Show MeSH
Related in: MedlinePlus