Characterizing the normal proteome of human ciliary body

Abstract

Background: The ciliary body is the circumferential muscular tissue located just behind the iris in the anterior chamber of the eye. It plays a pivotal role in the production of aqueous humor, maintenance of the lens zonules and accommodation by changing the shape of the crystalline lens. The ciliary body is the major target of drugs against glaucoma as its inhibition leads to a drop in intraocular pressure. A molecular study of the ciliary body could provide a better understanding about the pathophysiological processes that occur in glaucoma. Thus far, no large-scale proteomic investigation has been reported for the human ciliary body.

Results: In this study, we have carried out an in-depth LC-MS/MS-based proteomic analysis of normal human ciliary body and have identified 2,815 proteins. We identified a number of proteins that were previously not described in the ciliary body including importin 5 (IPO5), atlastin-2 (ATL2), B-cell receptor associated protein 29 (BCAP29), basigin (BSG), calpain-1 (CAPN1), copine 6 (CPNE6), fibulin 1 (FBLN1) and galectin 1 (LGALS1). We compared the plasma proteome with the ciliary body proteome and found that the large majority of proteins in the ciliary body were also detectable in the plasma while 896 proteins were unique to the ciliary body. We also classified proteins using pathway enrichment analysis and found most of proteins associated with ubiquitin pathway, EIF2 signaling, glycolysis and gluconeogenesis.

Conclusions: More than 95% of the identified proteins have not been previously described in the ciliary body proteome. This is the largest catalogue of proteins reported thus far in the ciliary body that should provide new insights into our understanding of the factors involved in maintaining the secretion of aqueous humor. The identification of these proteins will aid in understanding various eye diseases of the anterior segment such as glaucoma and presbyopia.

Figure 1

Schematic structure of the eye and experimental strategy for proteomic analysis of human ciliary body. Panel A. shows anatomy of the eye with a zoomed in view of the ciliary body. Panel B. depicts the proteomic workflow employed for the study.

Figure 2

MS/MS spectra of previously described proteins. A. The peptide IDLSNNLISSIDNDAFR from opticin B. shows the MS/MS spectra of peptide TEAPSATGQASSLLGGR from collagen, type XVIII, alpha 1 C. The Peptide QVLEGHVLSEAR belongs to cytochrome p450 1B1 D. VWTSGQVEEYDLDADDINSR peptide from aquaporin 1.

Figure 3

MS/MS spectra of novel proteins identified. A. shows the MS/MS spectra of peptide, EEAENNLAAFR, from Desmin B. The peptide, GAEILEVLHSLPAVR, derived from 26S proteasome non-ATPase regulatory subunit 6 C. NVDILKDPETVK Peptide from exportin-1 D. SEDPDQQYLILNTAR Peptide from vacuolar sorting-associated protein 35.

Figure 4

Comparison of the ciliary body proteome with the aqueous humor and plasma proteome. Panel A shows comparison of the ciliary body proteins with plasma proteins annotated in the Plasma Proteome Database. Panel B depicts comparison of the ciliary body proteome with aqueous humor proteome annotated from the published literature. Panel C shows a comparison of proteins that are common to the ciliary body and plasma with those that are common to the ciliary body and the aqueous humor.

Figure 5

Subcellular localization and functional annotation of proteins identified from the ciliary body. A. Gene Ontology analysis for subcellular localization of identified proteins B. Molecular function of identified proteins C. Biological processes of the identified proteins. The data regarding proteins was obtained from Human Protein Reference Database (http://www.hprd.org).