The availability of human genome sequence has transformed biomedical research over the past decade. However, an equivalent map for the human proteome with direct measurements of proteins and peptides does not exist yet. Here, we present a draft map of the human proteome using high resolution Fourier transform mass spectrometry. In-depth proteomic profiling of 30 histologically normal human samples including 17 adult tissues, 7 fetal tissues and 6 purified primary hematopoietic cells resulted in identification of proteins encoded by 17,294 genes accounting for ~84% of the total annotated protein-coding genes in humans. A unique and comprehensive strategy for proteogenomic analysis enabled us to discover a number of novel protein-coding regions, which includes translated pseudogenes, non-coding RNAs and upstream ORFs. This large human proteome catalog (available as an interactive web-based resource at http://www.humanproteomemap.org) will complement available human genome and transcriptome data to accelerate biomedical research in health and disease.

Autosomal recessive childhood-onset severe retinal dystrophy (arCSRD) designates a heterogeneous group of disorders affecting rod and cone photoreceptors simultaneously. The most severe cases are termed Leber congenital amaurosis (LCA), while the less aggressive forms are usually considered juvenile retinitis pigmentosa. Recently, mutations in the retinal-specific guanylate cyclase gene were found in patients with LCA. Disease genes implicated in other forms of arCSRD are expected to encode proteins present in the neuroretina or in the retinal pigment epithelium (RPE). The RPE, a monolayer of cells separating the vascular-rich choroid and the neuroretina, is in intimate contact with the outer segments of rods and cones via the microvilli surrounding the photoreceptors. The RPE expresses a tissue-specific and evolutionarily highly conserved 61 kD protein (RPE65) present at high levels in vivo. Although the function of RPE65 is not yet known, an important role in the RPE/photoreceptor vitamin-A cycle is suggested by the fact that RPE65 associates both with serum retinol-binding protein and with the RPE-specific 11-cis retinol dehydrogenase, an enzyme active in the synthesis of the visual pigment chromophore 11-cis retinal. Here we report that the analysis of RPE65 in a collection of about 100 unselected retinal-dystrophy patients of different ethnic origin revealed five that are likely to be pathogenic mutations, including a missense mutation (Pro363Thr), two point mutations affecting splicing (912 + 1G-->T and 65 + 5G-->A) and two small re-arrangements (ins144T and 831del8) on a total of nine alleles of five patients with arCSRD. In contrast to other genes whose defects have been implicated in degenerative retinopathies, RPE65 is the first disease gene in this group of inherited disorders that is expressed exclusively in the RPE, and may play a role in vitamin-A metabolism of the retina.

BackgroundThe vitreous humor is a transparent, gelatinous mass whose main constituent is water. It plays an important role in providing metabolic nutrient requirements of the lens, coordinating eye growth and providing support to the retina. It is in close proximity to the retina and reflects many of the changes occurring in this tissue. The biochemical changes occurring in the vitreous could provide a better understanding about the pathophysiological processes that occur in vitreoretinopathy. In this study, we investigated the proteome of normal human vitreous humor using high resolution Fourier transform mass spectrometry.ResultsThe vitreous humor was subjected to multiple fractionation techniques followed by LC-MS/MS analysis. We identified 1,205 proteins, 682 of which have not been described previously in the vitreous humor. Most proteins were localized to the extracellular space (24%), cytoplasm (20%) or plasma membrane (14%). Classification based on molecular function showed that 27% had catalytic activity, 10% structural activity, 10% binding activity, 4% cell and 4% transporter activity. Categorization for biological processes showed 28% participate in metabolism, 20% in cell communication and 13% in cell growth. The data have been deposited to the ProteomeXchange with identifier PXD000957.ConclusionThis large catalog of vitreous proteins should facilitate biomedical research into pathological conditions of the eye including diabetic retinopathy, retinal detachment and cataract.

The aqueous humor is a colorless, transparent fluid that fills the anterior chamber of the eye. It plays an important role in maintaining the intraocular pressure and providing nourishment to the lens and cornea. The constitution of the aqueous humor is controlled by the blood-aqueous barrier. Though this ocular fluid has been extensively studied, its role in ocular physiology is still not completely understood. In this study, aqueous humor samples were collected from 250 patients undergoing cataract surgery, subjected to multiple fractionation strategies and analyzed on a Fourier transform LTQ-Orbitrap Velos mass spectrometer. In all, we identified 763 proteins, of which 386 have been identified for the first time in this study. Sorbitol dehydrogenase (SORD), filensin (BFSP1), and phakinin (BFSP2) are some of the proteins that have not been previously reported in the aqueous humor. Gene Ontology analysis revealed 35% of the identified proteins to be extracellular, with a majority of them involved in cell communication and signal transduction. This study comprehensively reports 386 novel proteins that have important potential as biomarker candidates for future research into personalized medicine and diagnostics aimed towards improving visual health.