Mainak Sengupta

Cytogenetic damage and genetic variants in the individuals susceptible to arsenic‐induced cancer through drinking water

In West Bengal, India, more than 300,000 arsenic‐exposed people are showing symptoms of arsenic toxicity, which include cancers of skin and different internal organs. Since only 15–20% of the exposed population manifest arsenic‐induced skin lesions, it is thought that genetic variation might play an important role in arsenic toxicity and carcinogenicity. A total of 422 unrelated arsenic‐exposed subjects (244 skin‐symptomatic and 178 asymptomatic) were recruited for this study. Cytogenetic damage, as measured by chromosomal aberrations in lymphocytes and micronuclei formation in oral mucosa cells, urothelial cells and binucleated lymphocytes, was studied in unexposed, skin‐symptomatic and asymptomatic individuals with similar socioeconomic status. Identification of null mutations in GSTT1 and GSTM1 genes were carried out by PCR amplification. GSTP1 SNPs, implicated in susceptibility to various cancers, were assessed by PCR‐RFLP method. Symptomatic individuals had higher level of cytogenetic damage compared to asymptomatic individuals and asymptomatic individuals had significantly higher genotoxicity than unexposed individuals. No difference in allelic variants in GSTT1 and GSTP1 was observed between these 2 groups. Incidence of GSTM1 null gene frequencies was significantly higher in the asymptomatic group. Individuals with GSTM1‐positive (at least one allele) had significantly higher risk of arsenic‐induced skin lesions (odds ratio, 1.73; 95% confidence interval, 1.24–2.22). These results show a protective role of GSTM1 null in arsenic toxicity. This study also indicates that asymptomatic individuals are sub clinically affected and are also significantly susceptible to arsenic‐induced genotoxicity.

Molecular basis of albinism in India: Evaluation of seven potential candidate genes and some new findings

Albinism represents a group of genetic disorders with a broad spectrum of hypopigmentary phenotypes dependent on the genetic background of the patients. Oculocutaneous albinism (OCA) patients have little or no pigment in their eyes, skin and hair, whereas ocular albinism (OA) primarily presents the ocular symptoms, and the skin and hair color may vary from near normal to very fair. Mutations in genes directly or indirectly regulating melanin production are responsible for different forms of albinism with overlapping clinical features. In this study, 27 albinistic individuals from 24 families were screened for causal variants by a PCR-sequencing based approach. TYR, OCA2, TYRP1, SLC45A2, SLC24A5, TYRP2 and SILV were selected as candidate genes. We identified 5 TYR and 3 OCA2 mutations, majority in homozygous state, in 8 unrelated patients including a case of autosomal recessive ocular albinism (AROA). A homozygous 4-nucleotide novel insertion in SLC24A5 was detected in a person showing with extreme cutaneous hypopigmentation. A potential causal variant was identified in the TYRP2 gene in a single patient. Haplotype analyses in the patients carrying homozygous mutations in the classical OCA genes suggested founder effect. This is the first report of an Indian AROA patient harboring a mutation in OCA2. Our results also reveal for the first time that mutations in SLC24A5 could contribute to extreme hypopigmentation in humans.

A novel COMMD1 mutation Thr174Met associated with elevated urinary copper and signs of enhanced apoptotic cell death in a Wilson Disease patient

Wilson disease (WD) results from accumulation of copper and caused due to mutations in ATP7B, a copper transporting ATPase. Besides regular hepatic and neurological symptoms, WD patients occasionally manifest atypical symptoms due to unknown cause. To understand the molecular etiology of atypical WD manifestations, we screened COMMD1, a gene implicated in canine copper toxicosis, in 109 WD patients including those with atypical symptoms. In a patient showing apoptotic symptoms and high urinary copper surpassing normal WD levels, we identified a novel, putative mutation in COMMD1. Two other changes were also identified in the gene. We have examined genotype-phenotype correlation between the detected changes and the atypical presentation of the WD patient.

OCA1 in Different Ethnic Groups of India is Primarily Due to Founder Mutations in the Tyrosinase Gene

Oculocutaneous albinism (OCA) is a heterogeneous group of autosomal recessive disorders characterized by an abnormally low amount of melanin in the eyes, skin and hair, and associated with common developmental abnormalities of the eye. Defects in the tyrosinase gene (TYR) cause a common type of OCA, known as oculocutaneous albinism type 1 (OCA1). The molecular basis of OCA has been studied extensively in different population groups, but very little information is available on Indian patients. Our investigation covering thirteen ethnic groups of India, some representing >20 million people, revealed that among 25 OCA families 12 were affected with OCA1, and that these cases were primarily due to founder mutations in TYR. We detected nine mutations and eight SNPs in TYR, of which six mutations (five point mutations & one gross deletion) were novel. In contrast to most reports describing compound heterozygotes, the presence of homozygotes in 10 out of the 12 pedigrees underscores the lack of intermixing between these ethnic groups in India. Haplotype analysis suggested a few founder chromosomes causing the disease in the majority of the patients. Direct detection of the mutations prevalent in specific ethnic groups could be used for carrier detection and genetic counselling.

Indian genetic disease database

Indians, representing about one-sixth of the world population, consist of several thousands of endogamous groups with strong potential for excess of recessive diseases. However, no database is available on Indian population with comprehensive information on the diseases common in the country. To address this issue, we present Indian Genetic Disease Database (IGDD) release 1.0 (http://www.igdd.iicb.res.in)—an integrated and curated repository of growing number of mutation data on common genetic diseases afflicting the Indian populations. Currently the database covers 52 diseases with information on 5760 individuals carrying the mutant alleles of causal genes. Information on locus heterogeneity, type of mutation, clinical and biochemical data, geographical location and common mutations are furnished based on published literature. The database is currently designed to work best with Internet Explorer 8 (optimal resolution 1440 × 900) and it can be searched based on disease of interest, causal gene, type of mutation and geographical location of the patients or carriers. Provisions have been made for deposition of new data and logistics for regular updation of the database. The IGDD web portal, planned to be made freely available, contains user-friendly interfaces and is expected to be highly useful to the geneticists, clinicians, biologists and patient support groups of various genetic diseases.

Comprehensive analysis of the molecular basis of oculocutaneous albinism in Indian patients lacking a mutation in the tyrosinase gene

Background  Oculocutaneous albinism (OCA) refers to a group of inherited disorders where the patients have little or no pigment in the eyes, skin and hair. Mutations in genes regulating multi‐step melanin biosynthesis are the basis of four ‘classical’ OCA types with overlapping clinical features. There are a few reports on defects in TYR and a single report on SLC45A2 in Indians affected with OCA but no report on OCA2 (a major locus related to the disease) and TYRP1.

Structure-Function Correlation Analysis of Connexin50 Missense Mutations Causing Congenital Cataract: Electrostatic Potential Alteration Could Determine Intracellular Trafficking Fate of Mutants

Connexin50 (Cx50) mutations are reported to cause congenital cataract probably through the disruption of intercellular transport in the lens. Cx50 mutants that undergo mistrafficking have generally been associated with failure to form functional gap junction channels; however, sometimes even properly trafficked mutants were found to undergo similar consequences. We hereby wanted to elucidate any structural bases of the varied functional consequences of Cx50 missense mutations through in silico approach. Computational studies have been done based on a Cx50 homology model to assess conservation, solvent accessibility, and 3-dimensional localization of mutated residues as well as mutation-induced changes in surface electrostatic potential, H-bonding, and steric clash. This was supplemented with meta-analysis of published literature on the functional properties of connexin missense mutations. Analyses revealed that the mutation-induced critical alterations of surface electrostatic potential in Cx50 mutants could determine their fate in intracellular trafficking. A similar pattern was observed in case of mutations involving corresponding conserved residues in other connexins also. Based on these results the trafficking fates of 10 uncharacterized Cx50 mutations have been predicted. Further experimental analyses are needed to validate the observed correlation.

SNPs in genes with copy number variation: A question of specificity

The specificity of single nucleotide polymorphisms (SNPs) is likely to be compromised with most of the current PCRbased methods used to genotype a target locus in the presence of a highly homologous duplicated region. Such a lack of locus specificity could inflate the heterozygosity of the SNPs. We reasoned that public database for SNPs might be influenced by false allele calls, specifically in genes with copy number variation (CNV). Therefore, we compared the fraction of SNPs with high heterozygosity values (≥ 0.4) in NCBI dbSNP for genes with and without CNVs. Our observation highlights the challenges of selecting SNPs in genes with CNV for usage in complex biological studies.

Functional assessment of tyrosinase variants identified in individuals with albinism is essential for unequivocal determination of genotype to phenotype correlation.

Oculocutaneous albinism type 1 (OCA1), caused by pathogenic variations in the tyrosinase gene (TYR), is the most frequent and severe form of hypopigmentary disorder worldwide. While OCA1A manifests as a complete loss of melanin pigment, patients with OCA1B show residual pigmentation of the skin, hair and eyes. Limited experimental evidence suggests retention of TYR in the endoplasmic reticulum (ER) causes OCA1 pathogenesis. However, a comprehensive functional analysis of TYR missense variations and correlation with genotype is lacking.

TYRP1 (tyrosinase-related protein 1)

TYRP1 gene, having a chromosomal location of 9p23, encodes a melanosomal enzyme belonging to the tyrosinase family. TYRP1 catalyses oxidation of 5,6-dihydroxyindole-2-carboxylic acid (DHICA) into indole-5,6-quinone-2-carboxylic acid. TYRP1 is also thought to play a role in stabilizing tyrosinase and modulates its catalytic activity, in maintenance of melanosome structure, affecting melanocyte proliferation and melanocyte cell death. Defects in this gene cause oculocutaneous albinism type III; OCA III (also known as rufous oculocutaneous albinism).