Tyrosinase (TYR) gene sequencing and literature review reveals recurrent mutations and multiple population founder gene mutations as causative of oculocutaneous albinism (OCA) in Pakistani families.

PURPOSE: To investigate eight previously unreported Pakistani families with genetically undefined OCA for mutations in TYR. METHODS: Sanger sequencing of TYR has been performed in eight families with OCA phenotype. Mutation analysis was performed to establish the pathogenic role of novel mutation. Bioinformatics analysis was performed to predict the structural and functional impacts on protein due to the mutation. RESULTS: In this study, we identified six likely pathogenic variants of TYR (c.272 G>A, c.308 G>A, c.346C>T, c.715 C>T, c.832 C>T and c.1255 G>A), including one novel variant (c.308 G>A; p.Cys103Tyr), segregating as appropriate in each family. Cys103 lies in the highly conserved region of the tyrosinase enzyme, and p.Cys103Tyr is predicted to disturb enzymatic function via alteration of the configurational orientation of TYR leading to a more rigid polypeptide structure. We have also reviewed the mutation spectrum of TYR in Pakistani ethnicity. Published data on OCA families proposed that ~40% have been associated with genetic variations in the TYR gene. The mutations reported in this study have now been described with varying frequencies in Pakistani families, including very rare/unique mutations. CONCLUSION: A literature review of TYR gene mutations in Pakistani populations, combined with our genetic data, identified a number of gene mutations likely to represent regional ancestral founder mutations of relevance to Pakistani populations, in addition to sporadic and recurrent 'hotspot' mutations present repeatedly in other regions worldwide.

Distribution of IL28B and IL10 polymorphisms as genetic predictors of treatment response in Pakistani HCV genotype 3 patients.

There are over 10 million hepatitis C virus (HCV)-infected patients in Pakistan. For these patients, a combination of interferon with ribavirin is the most economical and easily available treatment. Single-nucleotide polymorphisms in interleukin genes have been reported to be associated with the pathogenesis and clearance of HCV, and sustained virologic response (SVR). An interleukin 28B (IL28B) gene polymorphism has been shown to modify treatment outcomes, but the effects of interleukin 10 (IL10) polymorphisms have not been previously assessed in the Pakistani population. The present study was conducted with 302 subjects categorized into two groups: 100 healthy volunteers (Group I) and 202 patients with chronic HCV (Group II). Patients within Group II were further divided into two subgroups according to therapeutic response: SVR (responders = 132) and NR (non-responders/relapsers = 70). IL28B (rs8099917, rs12979860) and IL10 (rs1800872, rs1800871, rs1800896) gene polymorphisms were studied in all subjects. A significant difference in the distribution of IL28B rs12979860C/T genotypes between the two groups (p<0.05) was observed, while of the three IL10 polymorphisms, a significant difference was only shown for rs1800896 A/G. Haplotype analysis (IL28B and IL10) showed a significant association of TTGTC and TTGTA when comparing the groups. There was a strong association of the favorable alleles rs8099917T and rs12979860C in the SVR group as compared with the NR group (p<0.05), and rs1800896 also showed an association with the SVR group as compared to the NR group (p<0.004). Haplotype analysis showed significant associations when comparing the SVR and NR subgroups, i.e. TCATC (p=0.009), TTGTA (p=0.005), TCATA (p<0.0005), TCACA (p=0.002), GTGCC (p=0.002) and TCGTC (p=0.005). IL28B (rs8099917 and rs12979860) and IL10 (rs1800896) polymorphisms alone, or in combination, are good predictors of therapeutic response in HCV-3a patients.

Metachromatic Leukodystrophy (MLD): a Pakistani Family with Novel ARSA Gene Mutation.

A deficiency of the enzyme arylsulfatase A (ARSA) causes a progressive neurodegenerative lysosomal storage disease known as metachromatic leukodystrophy (MLD). Diagnosis is based on the onset of neurological symptoms, presence of gait abnormalities, spasticity, decreased muscle stretch reflexes and neuro-radiological evidence of demyelination. The purpose of the present study was to identify any mutation in the candidate ARSA gene in a family of late infantile MLD patients. The diagnosis of suspected MLD patients was confirmed by a MRI report and low ARSA enzymatic activity in leukocytes. Sanger sequencing of full-length coding regions of ARSA gene was performed. Changes in the nucleotide sequence were determined by comparing the obtained data with the wild-type sequence. mRNA expression was analysed using real-time PCR. A novel base pair substitution at position c.338T>C (p.L113P) of ARSA gene was observed in the family and was confirmed in a normal population via ARMS-PCR and Sanger sequencing. The mRNA expression of ARSA gene showed a significant difference between normal and carrier individuals (p = 0.0008). In silico analysis by POLYPHEN, a pathogenicity prediction tool, predicted the possible damaging nature of this mutation. I-TASSER, a protein-modelling server, demonstrated the effects of this mutation on different domains of the ARSA protein, which plays a crucial role in the structural and functional integrity of enzyme. The novel p.L113P mutation in a Pakistani family with late infantile MLD has a pathogenic and destructive effect on the protein structure and function of ARSA. It is the first case reported in a Pakistani population using genetic analysis.

Hepatitis C virus entry: role of host and viral factors.

Hepatitis C virus (HCV) has been considered to be a significant risk factor in developing liver associated diseases including hepatocellular carcinoma all over the world. HCV is an enveloped positive strand virus comprising a complex between genomic RNA and viral envelope glycoproteins (E1 and E2), which are anchored within host derived double-layered lipid membrane surrounding the nucleocapsid composed of several copies of core protein. HCV cell entry is the first step in infection and viral replication into host cells mainly hepatocytes. HCV cell entry is a complex process involving both the viral (envelope glycoproteins E1/E2) and host factors (cellular receptors and associated factors i.e. CD81, SR-BI, LDL-R, CLDN1, Occludin, DC-SIGN, L-SIGN and Glycosaminoglycans). Besides these the expression of certain other conditions such as polarization and EWI-2 expression inhibits the viral cell entry. Exploring the mechanism of HCV entry will help to better understand the viral life cycle and possible therapeutic targets against HCV infection including viral and host factors involved in this process. New strategies such as RNAi represents a new option for targeting the host or viral factors for prevention and therapeutic against HCV infection. In the current review we try to summarize the current knowledge about mechanism and interaction of cellular and viral factors involved in HCV cell entry and its implication as therapeutic target to inhibit HCV infection.