The retina proteome Retina transcriptome Protein expression of genes elevated in retina Gene expression shared between retina and other tissues Proteins analyzed in retina Retina function Background Relevant links and publications

The retina proteome

The retina is a multilayered neural tissue located in the eye and converts light into nerve signals that are transported to the visual centers in the brain by the optical nerve. The retina originates from neuroepithelium (an outpocketing of the diencephalon) and consists of a network of neuronal and glial cells connected via synapses. Consequently, it shares the developmental origin and organization with other parts of the central nervous system (CNS). Transcriptome analysis shows that 79% (n=15630) of all human proteins (n=19670) are expressed in the retina and 310 of these genes show an elevated expression in the retina compared to other tissue types.

310 elevated genes
87 enriched genes
79 group enriched genes
Retina has most group enriched gene expression in common with brain

Figure 1. Schematic overview of the organization of the retina. Pigment epithelium cells is located nearest the choroid, while the nerve fiber layer is closest to the vitreous humour of the eye.

Retina transcriptome

Transcriptome analysis of the retina can be visualized with regard to specificity and distribution of transcribed mRNA molecules (Figure 1). Specificity illustrates the number of genes with elevated or non-elevated expression in the retina compared to other tissues. Elevated expression includes three subcategory types of elevated expression:

Tissue enriched: At least four-fold higher mRNA level in retina compared to any other tissues.
Group enriched: At least four-fold higher average mRNA level in a group of 2-5 tissues compared to any other tissue.
Tissue enhanced: At least four-fold higher mRNA level in retina compared to the average level in all other tissues.

Distribution, on the other hand, visualizes how many genes that have, or do not have, detectable levels (NX≥1) of transcribed mRNA molecules in the retina compared to other tissues. As evident in Table 1, all genes elevated in retina are categorized as:

Detected in single: Detected in a single tissue
Detected in some: Detected in more than one but less than one third of tissues
Detected in many: Detected in at least a third but not all tissues
Detected in all: Detected in all tissues

A. Specificity

B. Distribution

Figure 2. (A) The distribution of all genes across the five categories based on transcript specificity in retina as well as in all other tissues. (B) The distribution of all genes across the six categories, based on transcript detection (NX≥1) in retina as well as in all other tissues.

As shown in Figure 1, 310 genes show some level of elevated expression in the retina compared to other tissues. The three categories of genes with elevated expression in retina compared to other organs are shown in Table 1. In Table 2, the 12 genes with the highest enrichment in retina are defined.

Table 1. Number of genes in the subdivided categories of elevated expression in retina.

		Distribution in the 37 tissues
		Detected in single	Detected in some	Detected in many	Detected in all	Total
Specificity	Tissue enriched	0	61	23	3	87
	Group enriched	0	48	30	1	79
	Tissue enhanced	0	34	101	9	144
	Total	0	143	154	13	310

Table 2. The 12 genes with the highest level of enriched expression in retina. "Tissue distribution" describes the transcript detection (NX≥1) in retina as well as in all other tissues. "mRNA (tissue)" shows the transcript level in retina as NX values. "Tissue specificity score (TS)" corresponds to the fold-change between the expression level in retina and the tissue with second highest expression level.

Gene	Description	Tissue distribution	mRNA (tissue)	Tissue specificity score
RHO	rhodopsin	Detected in some	415.0	151
OPTC	opticin	Detected in some	245.8	92
RCVRN	recoverin	Detected in some	208.6	75
RBP3	retinol binding protein 3	Detected in some	135.2	45
CRX	cone-rod homeobox	Detected in some	125.2	42
GNAT1	G protein subunit alpha transducin 1	Detected in some	171.9	39
AIPL1	aryl hydrocarbon receptor interacting protein like 1	Detected in some	123.7	37
AC010319.2		Detected in some	41.6	32
TSPAN10	tetraspanin 10	Detected in some	117.4	29
SAG	S-antigen visual arrestin	Detected in some	215.8	26
NR2E3	nuclear receptor subfamily 2 group E member 3	Detected in some	50.2	26
RPE65	RPE65, retinoid isomerohydrolase	Detected in some	268.3	25

Protein expression of genes elevated in retina

In-depth analysis of the elevated genes in retina using antibody-based protein profiling allowed us to understand the distribution of the retina specific genes and their expression profiles. Proteins expressed in specific cell-types or in synapses were identified among the genes with elevated expression in the retina.

Proteins specifically expressed in pigment epithelial cells

Pigment epitheli cells produce melanin to protect photoreceptor cells from UV-radiation. Our analysis revealed that, in addition to melanocytes in the skin, melan-A (MLANA) is also expressed in pigment epitheli cells. Pigment epitheli cells are also involved in the transport of nutrients and ions between photoreceptor cells and blood vessels in the choroid. Another protein identified in pigment epitheli cells is bestrophin 1 (BEST1); it is known to form a bicarbonate transporting channel. Solute carrier family 16 member 8 (SLC16A8) plays a role in the transport of molecules and here we show that it is located in the cell membrane of pigment epitheli cells.

MLANA
BEST1
SLC16A8

Proteins specifically expressed in photoreceptor cells

An example of a protein located in the nuclei of photoreceptor cells is cone-rod homeobox (CRX), a transcription factor essential for normal photoreceptor function. The process of conversion of light to nerve signals, also called phototransduction, involves interaction between numerous proteins in the outer segments of photoreceptor cells,. There are two types of photoreceptor cells: rods and cones. In rods, phototransduction begins when light activates rhodopsin (RHO), followed by interaction with the rod-specific transducin. A component of the latter is G protein subunit gamma transducin (GNGT1). Another protein expressed in the outer segments is cyclic GMP-phosphodiesterase 6G (PDE6G). It converts cyclic GMP into 5' GMP in rods, a process known to lead to hyperpolarization and subsequent nerve signaling. S-antigen visual arrestin (SAG) turns off phototransduction by preventing interaction between rhodopsin and transducin. This allows activated rods to recover and consequently adapt to low light conditions.

CRX
RHO
GNGT1

PDE6G
SAG

Proteins involved in phototransduction in cones include g protein subunit gamma transducin 2 (GNGT2), guanylate cyclase activator 1B (GUCA1B) and arrestin 3 (ARR3). GNGT2 is a component of the cone-specific transducin, and like its rod-specific relative GNGT1, it is located in the outer segments. GUCA1B is a calcium-binding protein that activates photoreceptor guanylate cyclases. ARR3 is the cone-specific equivalent of SAG, and it is involved in the deactivation of cones.

GNGT2
GUCA1B
ARR3

Proteins specifically expressed in the outer limiting membrane

The outer limiting membrane is a row of cell-junctions located where the inner and outer segments of photoreceptors connect, as illustrated by integrin subunit beta 8 (ITGB8), which is a component of the integrin complex. Integrin complexes are important for cell to cell and cell to matrix interactions. Crumbs 2 (CRB2) is another protein expressed in the outer limiting membrane. CRB2 is a component of the Crumbs cell polarity complex, a protein important for many cellular processes during embryonic development. Mutation in this gene has been reported to cause congenital disorders such as retinitis pigmentosa.

ITGB8
CRB2

Proteins specifically expressed in the inner nuclear layer and ganglion cells

A transcription factor that may be involved in the development of cells in the inner nuclear layer (INL) is a visual system homeobox 2 (VSX2). VSX2 is expressed in the nuclei of a subset of cells in the INL. Recoverin (RCVRN), similarly to SAG, is known to block phototransduction in rods in darkness. Here we show that RCVRN is expressed in photoreceptor cells and in a subset of cells in INL. Calcium binding protein 5 (CABP5), is a member of a protein family involved in intracellular signaling. However, the exact function of this gene is unknown. A mouse homolog has been found in the INL of mice. Our analysis shows that CABP5 is expressed in the INL and ganglion cells in the human retina. The axons of ganglion cells are collected in the nerve fiber layer, as exemplified by PI16, and extend through the optic nerve to various visual centers in the central nervous system. PI16 is a peptidase inhibitor that has been reported to be involved in stress response in cardiac muscle and vasculature. Its function in the retina is not yet characterized.

VSX2
RCVRN

CABP5
PI16

Proteins specifically expressed in the inner and outer plexiform layers

The outer and inner plexiform layers (OPL and IPL) contain synapses through which the nerve signals are transmitted between the different cells in the retina. Anoctamin 2 (ANO2) encodes a calcium-activated chloride channel expressed exclusively in the OPL. CPLX4 is a protein located in the IPL and may be involved in synaptic vesicle exocytosis. Glutamate metabotropic receptor 6 (GRM6) is, as the name suggests, one of several types of metabotropic receptors that bind the neurotransmitter L-glutamate in the central nervous system. In the retina, GRM6 is detected in both the OPL and IPL.

ANO2
CPLX4
GRM6

Gene expression shared between retina and other tissues

There are 79 group enriched genes expressed in the retina. Group enriched genes are defined as genes showing a 4-fold higher average level of mRNA expression in a group of 2-5 tissues, including retina, compared to all other tissues.

In order to illustrate the relation of retina to other tissue types, a network plot was generated, displaying the number of genes shared between different tissue types (Figure 3).

Figure 3. An interactive network plot of the retina enriched and group enriched genes connected to their respective enriched tissues (grey circles). Red nodes represent the number of retina enriched genes and orange nodes represent the number of genes that are group enriched. The sizes of the red and orange nodes are related to the number of genes displayed within the node. Each node is clickable and results in a list of all enriched genes connected to the highlighted edges. The network is limited to group enriched genes in combinations of up to 3 tissues, but the resulting lists show the complete set of group enriched genes in the particular tissue.

Retina shares most group enriched gene expression with the brain (n=37), which is expected based on that retina and brain share developmental origin and organization. GNG13 is a protein expressed exclusively in synapses in the retina and brain, as well as a subset of cells in the INL of the retina. It is a gamma subunit of a heterotrimeric G protein, which is known to be expressed in taste, retinal and neuronal tissues.

GNG13 - retina
GNG13 - cerebellum

Skin is another tissue that shares expression of several (n=6) group enriched genes with the retina. Tyrosinkinase (TYR) converts tyrosine to melanin. It is expressed in melanocytes in skin and hair, and in pigment epithelial cells in the retina.

TYR - retina
TYR - hair

Proteins analyzed in retina

Protein expression of selected genes with RNA expression elevated in retina or literature indicating function related to retina were analyzed in retina samples. The full list of genes used for protein profiling in the retina is found in Table 3.

Table 3. Following 108 genes have been analyzed in retina.

Gene	Gene description	Staining pattern
ABCA4	ATP binding cassette subfamily A member 4	Staining in photoreceptor cells.
ABCG4	ATP binding cassette subfamily G member 4	Staining in photoreceptor, plexiform and nerve fiber layers.
AC010319.2		Staining mainly in outer limiting membrane.
AIPL1	Aryl hydrocarbon receptor interacting protein like 1	Positivity in photoreceptor cells.
ANKRD33	Ankyrin repeat domain 33	Staining in photoreceptors.
ANO2	Anoctamin 2	Positivity in outer plexiform layer.
ARMS2	Age-related maculopathy susceptibility 2	Positivity in outer and inner plexiform layers.
ARR3	Arrestin 3	Staining in cones.
ATP1B2	ATPase Na+/K+ transporting subunit beta 2	Staining in most layers, strongest in photoreceptor cells and inner nuclear layer.
BCO2	Beta-carotene oxygenase 2	Staining in photoreceptor cells and pigment epithelium.
BEST1	Bestrophin 1	Positivity in basal membrane of retinal pigment epithelium.
CABP5	Calcium binding protein 5	Positivity in ganglion cells and cells in inner nuclear layer.
CACNA1F	Calcium voltage-gated channel subunit alpha1 F	Positivity in photoreceptor cells segments.
CDHR1	Cadherin related family member 1	Positivity in photoreceptor cell outer segments.
CFAP161	Cilia and flagella associated protein 161	Staining in cones and nerve fibers.
CHRNA3	Cholinergic receptor nicotinic alpha 3 subunit	Positivity in cells in photoreceptor layer, nerve fibers in outer and inner plexiform layer and nerve fiber layer.
CLIC6	Chloride intracellular channel 6	Positivity in nerve fibers in nerve fiber layer.
CLUL1	Clusterin like 1	Positivity in rods.
CNGB3	Cyclic nucleotide gated channel beta 3	Positivity in outer liming membrane.
COL9A1	Collagen type IX alpha 1 chain	Positivity in cones.
CPLX4	Complexin 4	Positivity in outer and inner plexiform layers.
CRABP1	Cellular retinoic acid binding protein 1	Positivity in inner nuclear layer and nerve fibers.
CRB1	Crumbs 1, cell polarity complex component	Positivity in photoreceptor cells and in outer plexiform layer.
CRB2	Crumbs 2, cell polarity complex component	Positivity in outer limiting membrane.
CRX	Cone-rod homeobox	Positivity in photoreceptor cells.
EFEMP1	EGF containing fibulin extracellular matrix protein 1	Positivity in rods, outer and inner plexiform layers and nerve fiber layer.
EGFLAM	EGF like, fibronectin type III and laminin G domains	Staining in outer plexiform layer.
FMN1	Formin 1	Staining in photoreceptor cells and outer plexiform layer.
FSCN2	Fascin actin-bundling protein 2, retinal	Positivity in photoreceptor cells.
GNAT1	G protein subunit alpha transducin 1	Positivity in rod segments.
GNG13	G protein subunit gamma 13	Membranous positivity in inner nuclear layer and plexiform layers.
GNGT1	G protein subunit gamma transducin 1	Positivity in rods.
GNGT2	G protein subunit gamma transducin 2	Positivity in cones.
GPR61	G protein-coupled receptor 61	Positivity in photoreceptor cell segments and outer plexiform layer.
GRK1	G protein-coupled receptor kinase 1	Positivity in photoreceptor cells.
GRK7	G protein-coupled receptor kinase 7	Cytoplasmic positivity in cells in photoreceptor layer, nerve fibers in outer plexiform layer and in nerve fiber layer.
GRM6	Glutamate metabotropic receptor 6	Staining in nerve fibers.
GUCA1A	Guanylate cyclase activator 1A	Staining mainly in photoreceptor layer, strongest in cones.
GUCA1B	Guanylate cyclase activator 1B	Positivity in cones and outer plexiform layer.
HCN1	Hyperpolarization activated cyclic nucleotide gated potassium channel 1	Cytoplasmic positivity in cells in photoreceptor layer, inner nuclear layer, inner and outer plexiform layer and nerve fiber layer.
IMPG1	Interphotoreceptor matrix proteoglycan 1	Positivity in photoreceptor cells.
IMPG2	Interphotoreceptor matrix proteoglycan 2	Positivity in photoreceptor cell segments
IQCB1	IQ motif containing B1	Positivity in cones.
ITGB8	Integrin subunit beta 8	Positivity in outer limiting membrane of retina.
KCNB1	Potassium voltage-gated channel subfamily B member 1	Staining in rods and subset of cells in inner nuclear layer.
KCNC1	Potassium voltage-gated channel subfamily C member 1	Positivity in nerve fibers in plexiform layers.
KCNV2	Potassium voltage-gated channel modifier subfamily V member 2	Positivity in photoreceptor cells.
LRIT2	Leucine rich repeat, Ig-like and transmembrane domains 2	Positivity in rods and nerve fibers.
LRIT3	Leucine rich repeat, Ig-like and transmembrane domains 3	Positivity in photoreceptor cell segments.
MGARP	Mitochondria localized glutamic acid rich protein	Staining mainly in nerve fibers, cells in inner nuclear layer and ganglion cells.
MLANA	Melan-A	Staining in pigment epithelium.
MPP1	Membrane palmitoylated protein 1	Staining in photoreceptor cells and outer plexiform layer.
MPP5	Membrane palmitoylated protein 5	Positivity in outer limiting membrane.
MSI1	Musashi RNA binding protein 1	Staining in inner nuclear and ganglion cell layers.
MTNR1B	Melatonin receptor 1B	Staining in nerve fiber layer.
MYO3A	Myosin IIIA	Staining in mainly in photoreceptor cells and nerve fibers.
NPVF	Neuropeptide VF precursor	Positivity in nerve fibers.
NRL	Neural retina leucine zipper	Positivity in rods.
OPN1SW	Opsin 1, short wave sensitive	Positivity in outer segments of rods.
OPN4	Opsin 4	Positivity in nerve fibers in outer and inner plexiform layers and nerve fiber layer.
OPN5	Opsin 5	Positivity in photoreceptor cells.
PAX6	Paired box 6	Staining in inner nuclear and ganglion cell layers.
PDC	Phosducin	Positivity in cells in photoreceptor layer.
PDE6A	Phosphodiesterase 6A	Positivity in rods.
PDE6B	Phosphodiesterase 6B	Positivity in outer segments of photoreceptor cells
PDE6G	Phosphodiesterase 6G	Positivity in outer segments of rods.
PDE6H	Phosphodiesterase 6H	Positivity in outer segments of rods.
PI16	Peptidase inhibitor 16	Positivity in nerve fiber layer.
PLA2G5	Phospholipase A2 group V	Positivity mainly in photoreceptor cells.
PLCB4	Phospholipase C beta 4	Positivity mainly in photoreceptor cells.
POU4F1	POU class 4 homeobox 1	Positivity in nerve fibers, subsets of cells in inner nuclear layer and pigment epithelium.
PRKCZ	Protein kinase C zeta	Staining in photoreceptor cells.
PRPH2	Peripherin 2	Positivity in photoreceptor cell outer segments.
RARB	Retinoic acid receptor beta	Nuclear positivity in cells in photoreceptor layer, inner nuclear layer and ganglion cells.
RB1	RB transcriptional corepressor 1	Positivity was observed in photoreceptor cells, cells in inner nuclear layer and ganglion cells.
RBP3	Retinol binding protein 3	Positivity in photoreceptor cells.
RCVRN	Recoverin	Positivity in photoreceptor cells and subsets of cells in inner nuclear layer.
RD3	Retinal degeneration 3	Positivity in cones.
RDH13	Retinol dehydrogenase 13	Positivity in inner segments of photoreceptor cells and nerve fibers in plexiform layers.
RGS9	Regulator of G protein signaling 9	Positivity in photoreceptor segments.
RGS9BP	Regulator of G protein signaling 9 binding protein	Positivity in rod segments
RHO	Rhodopsin	Positivity in rod segments.
RLBP1	Retinaldehyde binding protein 1	Positivity in pigment epithelium, Mueller glia and inner nuclear layer.
ROM1	Retinal outer segment membrane protein 1	Positivity in photoreceptor layer.
RP1	RP1, axonemal microtubule associated	Positivity in cones and in outer plexiform layer.
RP1L1	RP1 like 1	Positivity in photoreceptor outer segments.
RPGRIP1	RPGR interacting protein 1	Positivity in segments of photoreceptor cells.
RS1	Retinoschisin 1	Positivity in photoreceptor cells and nerve fibers.
SAG	S-antigen visual arrestin	Positivity in rod segments.
SCUBE1	Signal peptide, CUB domain and EGF like domain containing 1	Cytoplasmic positivity in photoreceptor layer, inner and outer plexiform layer and nerve fiber layer.
SEMA4F	Ssemaphorin 4F	Positivity in nerve fiber layer.
SIX6	SIX homeobox 6	Positivity in photoreceptor layer and in subset of cells in inner nuclear layer.
SLC13A4	Solute carrier family 13 member 4	Positivity in nerve fiber layer.
SLC16A8	Solute carrier family 16 member 8	Positivity in pigment epithelium.
SLC1A7	Solute carrier family 1 member 7	Positivity in plexiform layers.
SLC24A1	Solute carrier family 24 member 1	Positivity in photoreceptor cell segments and subset of nerve fibers.
SLC32A1	Solute carrier family 32 member 1	Positivity n inner plexiform layer.
SLC6A6	Solute carrier family 6 member 6	Cytoplasmic and nuclear staining in all layers.
SPATA7	Spermatogenesis associated 7	Staining mainly in photoreceptor cell segments.
SPTBN5	Spectrin beta, non-erythrocytic 5	Staining in photoreceptor and inner nuclear layers.
TFAP2A	Transcription factor AP-2 alpha	Positivity in subsets of cells in inner nuclear and ganglion cell layers.
TFAP2B	Transcription factor AP-2 beta	Nuclear positivity in inner nuclear layer and ganglion cells.
TMEM221	Transmembrane protein 221	Staining in outer limiting membrane.
TRPM1	Transient receptor potential cation channel subfamily M member 1	Positivity in subset of cells in inner nuclear layer and in nerve fibers in outer plexiform layer.
TYR	Tyrosinase	Strong staining in pigment epithelium.
UNC119	Unc-119 lipid binding chaperone	Positivity in photoreceptor cells and outer plexiform layer.
VAT1L	Vesicle amine transport 1 like	Staining in photoreceptor layer, inner nuclear layer and nerve fibers.
VSX2	Visual system homeobox 2	Positivity in inner nuclear layer.
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Retina function

The retina is a multilayered neural tissue that comprises the innermost layer of the eye, called the inner photosensitive layer. Similar to other parts of the central nervous system, it originates from the neuroepithelium, an outpocketing of the diencephalon. In the retina, light is converted into nerve signals that are transported to the visual centers in the brain by the optical nerve. As light enters the eye through the pupil and becomes refracted by the lens, it reaches the retina where it is absorbed. Photosensitive neuronal cells called photoreceptor cells transduce the light into nerve signals that are then transmitted to different types of interneurons that modulate the nerve signaling before it is collected in the optical nerve.

There are two types of photoreceptor cells: rods and cones. Rod photoreceptors cells register the presence of light and allow vision during low light conditions while cones register red, green and blue colors and allow color vision. A layer of pigment epithelial cells provide nutrients and protection against UV-radiation to the photoreceptor cells. Four types of interneurons modulate nerve signaling by inhibiting and/or activating the cells they interact with, all located in the inner nuclear layer. Horizontal cells connect only to synapses of the photoreceptor cells, while bipolar cells transmit signals between photoreceptor cells and ganglion cells. Amacrine cells modulate signals coming from bipolar cells to ganglion cells. Ganglion cells are the last cells to receive and regulate molecular signals from the photoreceptor cells before nerve signals are collected in the optical nerve. They interact both with bipolar and amacrine cells. Muller glia cells, as the name suggests are glial cells. They maintain the structural integrity of the retina by stretching across all layers as well as buffer potassium and neurotransmitters.

Background

Here, the protein-coding genes expressed in retina are described and characterized, together with examples of immunohistochemically stained tissue sections that visualize corresponding protein expression patterns of genes with elevated expression in retina.

Transcript profiling was based on the FANTOM5 transcriptomics dataset, corresponding to a total of 77 mixed patient samples from 60 different human normal tissue types. The final consensus normalized expression (NX) value for each tissue type was used for classification of all genes according to the tissue specific expression into two different categories, based on specificity or distribution.

Relevant links and publications

Uhlén M et al., Tissue-based map of the human proteome. Science (2015)
PubMed: 25613900 DOI: 10.1126/science.1260419

Yu NY et al., Complementing tissue characterization by integrating transcriptome profiling from the Human Protein Atlas and from the FANTOM5 consortium. Nucleic Acids Res. (2015)
PubMed: 26117540 DOI: 10.1093/nar/gkv608

Fagerberg L et al., Analysis of the human tissue-specific expression by genome-wide integration of transcriptomics and antibody-based proteomics. Mol Cell Proteomics. (2014)
PubMed: 24309898 DOI: 10.1074/mcp.M113.035600