The proteome of human retina

Zhang, Pingbo; Dufresne, Craig; Turner, Randi; Ferri, Sara; Venkatraman, Vidya; Karani, Rabia; Lutty, Gerard A.; Eyk, Jennifer E. Van; Semba, Richard D.

doi:10.1002/pmic.201400397

Cited by 30 publications

(34 citation statements)

References 15 publications

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“…We tried to consider this dynamic behavior as far as possible and included new samples in the study during the development of the project. Consequently, we included 4 samples from rare biological sources, since it had been proved that these samples can be used to detect missing proteins: 13 spermatozoid, 13 seminal plasma, 26 retina, 27 and placenta. 28 In addition to that, we included a most recent proteome characterization of the HEK293 cell line 29 in replacement of the experiment with PRIDE accession number PXD001383.…”

Section: Resultsmentioning

confidence: 99%

Detection of Missing Proteins Using the PRIDE Database as a Source of Mass Spectrometry Evidence

et al. 2016

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The current catalogue of the human proteome is not yet complete, as experimental proteomics evidence is still elusive for a group of proteins known as the missing proteins. The Human Proteome Project (HPP) has been successfully using technology and bioinformatic resources to improve the characterization of such challenging proteins. In this manuscript, we propose a pipeline starting with the mining of the PRIDE database to select a group of data sets potentially enriched in missing proteins that are subsequently analyzed for protein identification with a method based on the statistical analysis of proteotypic peptides. Spermatozoa and the HEK293 cell line were found to be a promising source of missing proteins and clearly merit further attention in future studies. After the analysis of the selected samples, we found 342 PSMs, suggesting the presence of 97 missing proteins in human spermatozoa or the HEK293 cell line, while only 36 missing proteins were potentially detected in the retina, frontal cortex, aorta thoracica, or placenta. The functional analysis of the missing proteins detected confirmed their tissue specificity, and the validation of a selected set of peptides using targeted proteomics (SRM/MRM assays) further supports the utility of the proposed pipeline. As illustrative examples, DNAH3 and TEPP in spermatozoa, and UNCX and ATAD3C in HEK293 cells were some of the more robust and remarkable identifications in this study. We provide evidence indicating the relevance to carefully analyze the ever-increasing MS/MS data available from PRIDE and other repositories as sources for missing proteins detection in specific biological matrices as revealed for HEK293 cells.

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Section: Resultsmentioning

confidence: 99%

Detection of Missing Proteins Using the PRIDE Database as a Source of Mass Spectrometry Evidence

et al. 2016

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“…Alternative splicing is a dynamically regulated process that contributes to the proteomic diversity and it is especially common for genes expressed in the nervous system, where the synthesis of protein variants determines properties of different types of neurons [24, 25]. It has been estimated that 15% of disease causing point-mutations affect pre-mRNA splicing and impaired splicing is also known to contribute to retinal degeneration [26].…”

Section: Discussionmentioning

confidence: 99%

EYS Is a Protein Associated with the Ciliary Axoneme in Rods and Cones

Alfano¹,

Kruczek²,

Shah³

et al. 2016

PLoS ONE

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PurposeMutations in the EYS gene are a common cause of autosomal recessive retinitis pigmentosa (arRP), yet the role of the EYS protein in humans is presently unclear. The aim of this study was to investigate the isoform structure, expression and potential function of EYS in the mammalian retina in order to better understand its involvement in the pathogenesis of arRP.MethodsTo achieve the objective, we examined the expression of mRNA transcripts of EYS isoforms in human tissues and cell lines by RT-PCR. We also investigated the localisation of EYS in cultured cells and retinal cryo-sections by confocal fluorescence microscopy and Western blot analysis.ResultsRT-PCR analysis confirmed that EYS has at least four isoforms. In addition to the previously reported EYS isoforms 1 and 4, we present the experimental validation of two smaller variants referred to as EYS isoforms 2 and 3. All four isoforms are expressed in the human retina and Y79 cells and the short variants were additionally detected in the testis. Immunofluorescent confocal microscopy and Western blot analysis revealed that all EYS isoforms preferentially localise to the cytoplasm of Y79 and HeLa cells. Moreover, an enrichment of the endogenous protein was observed near the centrosomes in Y79 cells. Interestingly, EYS was observed at the ciliary axoneme in Y79 ciliated cells. In macaque retinal cryosections, EYS was found to localise in the region of the photoreceptor ciliary axoneme in both rods and cones as well as in the cytoplasm of the ganglion cells.ConclusionThe results obtained in this study lead us to speculate that, in photoreceptor cells, EYS could be a protein involved in maintaining the stability of the ciliary axoneme in both rods and cones. The variability of its isoform structure suggests that other roles are also possible and yet to be established.

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“…The first data set is a proteomic analysis on retinal tissue by Zhang et al 50 (http://proteomecentral.proteomexchange.org/cgi/GetDataset?ID=PXD001242). According to this study, proteins were extracted from five normal retinal tissues and fractionated using SDS-PAGE.…”

Section: Methodsmentioning

confidence: 99%

Functional Networks of Highest-Connected Splice Isoforms: From The Chromosome 17 Human Proteome Project

Menon

Govindarajoo

et al. 2015

J. Proteome Res.

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Alternative splicing allows a single gene to produce multiple transcript-level splice isoforms from which the translated proteins may show differences in their expression and function. Identifying the major functional or canonical isoform is important for understanding gene and protein functions. Identification and characterization of splice isoforms is a stated goal of the HUPO Human Proteome Project and of neXtProt. Multiple efforts have catalogued splice isoforms as “dominant”, “principal”, or “major” isoforms based on expression or evolutionary traits. In contrast, we recently proposed highest connected isoforms (HCIs) as a new class of canonical isoforms that have the strongest interactions in a functional network and revealed their significantly higher (differential) transcript-level expression compared to nonhighest connected isoforms (NCIs) regardless of tissues/cell lines in the mouse. HCIs and their expression behavior in the human remain unexplored. Here we identified HCIs for 6157 multi-isoform genes using a human isoform network that we constructed by integrating a large compendium of heterogeneous genomic data. We present examples for pairs of transcript isoforms of ABCC3, RBM34, ERBB2, and ANXA7. We found that functional networks of isoforms of the same gene can show large differences. Interestingly, differential expression between HCIs and NCIs was also observed in the human on an independent set of 940 RNA-seq samples across multiple tissues, including heart, kidney, and liver. Using proteomic data from normal human retina and placenta, we showed that HCIs are a promising indicator of expressed protein isoforms exemplified by NUDFB6 and M6PR. Furthermore, we found that a significant percentage (20%, p = 0.0003) of human and mouse HCIs are homologues, suggesting their conservation between species. Our identified HCIs expand the repertoire of canonical isoforms and are expected to facilitate studying main protein products, understanding gene regulation, and possibly evolution. The network is available through our web server as a rich resource for investigating isoform functional relationships (http://guanlab.ccmb.med.umich.edu/hisonet). All MS/MS data were available at ProteomeXchange Web site (http://www.proteomexchange.org) through their identifiers (retina: PXD001242, placenta: PXD000754).

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The proteome of human retina

Cited by 30 publications

References 15 publications

Detection of Missing Proteins Using the PRIDE Database as a Source of Mass Spectrometry Evidence

Detection of Missing Proteins Using the PRIDE Database as a Source of Mass Spectrometry Evidence

EYS Is a Protein Associated with the Ciliary Axoneme in Rods and Cones

Functional Networks of Highest-Connected Splice Isoforms: From The Chromosome 17 Human Proteome Project

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