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PURPOSE: The aim of this study was to determine if there is a common founder for the Thr377Met myocilin mutation in primary open angle glaucoma (POAG) families with various ethnic backgrounds. METHODS: Genomic DNA of 24 POAG-affected individuals from nine pedigrees with the Thr377Met mutation and 104 unaffected family members was genotyped with six microsatellite markers and four single nucleotide polymorphisms. The families were from Greece, India, Finland, the USA, and Australia. To assess the degree of linkage disequilibrium across MYOC in the general population we also investigated data generated from the HapMap consortium. RESULTS: Three distinct haplotypes associated with the Thr377Met myocilin mutation were identified. The families from the USA and Greece, as well as the three Australian families originating from Greece and the former Yugoslavian Republic of Macedonia had one common haplotype. Interestingly, however, HapMap data suggest that linkage disequilibrium across MYOC was not strong. CONCLUSIONS: The Thr377Met myocilin mutation has arisen at least three separate times. Evidence for genetic founder effects in this prevalent age-related, yet heterogeneous, disease has important implications for future gene identification strategies. Mol Vis

Purpose: In Bornholm eye disease, a defect in the splicing of transcripts from a variant OPN1LW opsin gene leads to a depletion in spliced transcript levels and, consequently, a reduction in photopigment in photoreceptors expressing the variant gene. Methods: Myopic and age-matched control subjects were drawn from the Western Australian Pregnancy Cohort (Raine) Study and the Norfolk Island Eye Study groups. The OPN1LW opsin gene was amplified using long-range PCR methodology and was fully sequenced. Expression of variant opsins was evaluated using quantitative PCR (qPCR). RNA secondary structure changes arising from identified variants were predicted by modeling. Results: Forty-two nucleotide sites were found to vary across the 111 subjects studied. Of these, 15 had not been previously reported, with three present only in myopic individuals. Expression of these variants in transfected human embryonic kidney (HEK293T) cells demonstrated that splicing efficiencies were not affected. However, gene transcripts from two of the three variants were significantly depleted. RNA secondary structure modeling predicted that these single nucleotide changes could affect RNA stability. Conclusions: None of the variants identified in myopic individuals appeared to alter the efficiency of transcript splicing. However, two resulted in a significant reduction in the number of spliced and unspliced transcripts, indicating an overall reduction in steady-state transcript stability. Such a change would be expected to result in a reduced amount of photopigment, and this may be a contributing factor in the development of myopia.

Purpose Pseudoexfoliation syndrome is a major risk factor for the development of glaucoma. Following recent reports of a strong association of coding variants in the lysyl oxidase-like 1 (LOXL1) gene with this syndrome but low penetrance and variable disease frequency between different populations, we aimed to identify additional genetic factors contributing to the disease. The clusterin (CLU) gene has been proposed as a candidate because of the presence of clusterin protein in pseudoexfoliation deposits, its varied levels in aqueous humor of cases compared to controls, and the role of the protein as a molecular chaperone. We investigated the association of genetic variants across CLU in pseudoexfoliation syndrome and analyzed molecular characteristics of the encoded protein in ocular tissues. Methods The expression of clusterin in relevant ocular tissues was assessed using western blotting. Nine tag single nucleotide polymorphisms (SNPs) across CLU were genotyped in 86 cases of pseudoexfoliation syndrome and 2422 controls from the Australian Blue Mountains Eye Study cohort. Each SNP and haplotype was assessed for association with the syndrome. Results Clusterin was identified in normal human iris, the ciliary body, lens capsule, optic nerve, and aqueous humor. Post-translational modification gives rise to a 100 kDa precursor protein in ocular tissues, larger than that reported in non-ocular tissues. One CLU SNP (rs3087554) was nominally associated with pseudoexfoliation syndrome at the genotypic level (p=0.044), although not when the age of controls was restricted to those over 73 years. Only age and the LOXL1 diplotype were significant factors in the logistic regression. One haplotype of all nine CLU SNPs was also associated (p=0.005), but the significance decreased slightly with the use of the age-restricted controls (p=0.011). Conclusions Clusterin is present in ocular anterior segment tissues involved in pseudoexfoliation syndrome. Although one haplotype may contribute in a minor way to genetic risk of pseudoexfoliation syndrome, common variation in this gene is not a major contributor to the risk of pseudoexfoliation syndrome.

Here we present an overview of our existing knowledge on the function of RIN4 as a regulator of plant defense and as a guardee of multiple plant R-proteins. Domain analysis of RIN4 reveals two NOI domains. The NOI domain was originally identified in a screen for nitrate induced genes. The domain is comprised of approximately 30 amino acids and contains 2 conserved motifs (PXFGXW and Y/FTXXF). The NOI gene family contains members exclusively from the plant lineage as far back as moss. In addition to the conserved NOI domain, members within the family also contain conserved C-terminal cysteine residue(s) which are sites for acylation and membrane tethering. Other than these two characteristic features, the sequence of the family of NOI-containing proteins is diverse and, with the exception of RIN4, their functions are not known. Recently published interactome data showing interactions between RIN4 and components of the exocyst complex prompt us to raise the hypothesis that RIN4 might be involved in defense associated vesicle trafficking.