The term “pigmentosa” deals with the characteristic appearance, during the advanced states of the disease, of abnormal areas of pigment into the retina. Degeneration involves both eyes and affects photoreceptors and Retinal Pigment Epithelium (RPE) [5], inducing a slow and progressive death in these cells and leading to lose ability to transmit brain the visual informations. Photoreceptors are particular light-sensitive cells: cones, so called for their characteristic shape, incorporate image and color details, and are mainly localized in the retina central part, called macula; rods, elongated and tapered, are involved in night vision and low light conditions, as well as in object motion perception, and are predominantly distributed in the peripheral zone [6]. Thanks to these receptors, for example, we can realize the danger that comes close to the corner of the eye, even if we are not able to perceive the details. Rods represent retinitis first and most involved photoreceptor system (cones are involved in advanced stages of disease, even if with a currently unknown mechanism): they undergo apoptosis, with a loss rate evidenced by ONL (Outer Nuclear layer, the retinal layer which contains them) decrease and retinal accumulation of pigment (rhodopsin) [7]. Generally, it can be assumed that the mere presence of the rods provides survival signals needed to keep “alive” cones.

About the clinical picture, the first symptom is (but not always) the decrease of crepuscular nocturnal visual acuity (difficulty in driving at night or move in dimly illuminated rooms), until you come to a narrowing of the peripheral visual field (difficulty in perceiving objects placed laterally or seeing the steps going down the stairs) and, in the final stage of the disease, the loss of central vision and blindness [8]. It can also be accompanied by cataract or deafness, events directly related to retinitis pigmentosa. Diagnosis [9] consists of fundus examination and visual field, followed by visus, electroretinogram and fluorescein angiography.

The disease progression rate and the age of symptom onset [10] vary according to many factors, including the pattern of genetic transmission. Today, it is known that a wide range of retinitis pigmentosa forms is caused by about 60 genes (Table 1), the most of them involved in phototransduction, canonical retinoid cycle in rods (twilight vision) and cones (daylight vision), inactivation, recovery and regulation of phototransduction cascade, cargo trafficking to the periciliary membrane and signal transduction [11]. A special mention goes to oxidative stress which, as for other hereditary pathologies [12], could determine the onset of retinitis pigmentosa [13].

Mutations of these genes may be autosomal recessive (50-60%), dominant (30-40%) [14]; [15], X-linked (5-20%; the first gene linked to the RP phenotype was discovered twenty-six years ago on the X chromosome) [16] or sporadic (30%). Although these known genes cause the most of retinitis pigmentosa forms, there are many other ones that are still unidentified nowadays [11, 17-27], and several regulative aspects have to be discovered [28]. This scenario suggests that new genes could contribute to the genesis of pathology, and should be investigated in order to improve treatment for this disabling disease. Our approach is based on an in-silico analysis.

Intersection of about 50 causative genes for retinitis pigmentosa by Cytoscape and its plugins gave us about 30 proximal correlated genes, from witch we extracted 5 ones that are involved into phototransduction pathway, specifically in dark adaptation phases, which result the most altered one in pathological retinitis phenotype [29] (Figs. 1-3).

In order to understand at what level the filtered 5 genes could be involved in retinal function alterations, it is necessary to explain briefly the main phases of phototransduction. The transduction of light into a neural signal takes place in the outer segment of photoreceptors: in our study we underline what happens in rods, which are retinitis pigmentosa most involved cells. Rod outer segment shows a lipid bilayer membrane arranged as flattened “discs” (optical discs), which is strictly packed with the photopigment rhodopsin. Rhodopsin is a member of the superfamily of seven-helix, G-Protein–Coupled Receptor proteins (GPCRs). Differently from chemosensing GPCRs, rhodopsin presents its ligand, the light-absorbing “chromophore” retinaldehyde, prebound. The bound form, 11-cis retinal, acts as a strong antagonist, holding rhodopsin in its completely inactive state. Absorption of a photon of light isomerizes the chromophore to the the all-trans configuration, which rapidly triggers a conformational change in the protein, activating rhodopsin as an enzyme. The conformational change triggered by light causes rhodopsin absorption spectrum to shift into the UV, so that the pigment loses its visible color and is said to “bleach.” Long after the end of signaling, the Schiff-base bond that attaches the all-trans retinal is hydrolyzed, permitting the retinoid to dissociate from the apo-protein, opsin. In addition to rhodopsin (R), which is packed into the disc membrane at high density, three other classes of proteins cooperate into activation of the photoresponse: 1) an heterotrimeric G - protein (G), called transducin in rods, with a 1:10 stechiometry to rhodopsin; 2) the cyclic nucleotide phosphodiesterase (PDE), which hydrolyzes cyclic GMP (cGMP), the second cytoplasmic messenger, present at about 100 units per rhodopsin one; and (3) the Cyclic Nucleotide – Gated Channels (CNGCs) of the plasma membrane, which control the flow of electrical current into the outer segment. The first three steps involve proteins associated with the disc membrane (rhodopsin, which is integral, and the G-protein and PDE, which are connected by acyl groups), whereas the final two involve the cGMP, and its action at the plasma membrane (Fig. 4 summarize all five steps).

Fig. (1). Intersection pathways of 50 known causative genes of retinitis pigmentosa. This image shows the Cytoscape pathways analysis, supported by GeneMANIA plug-in, with nodes and edges reflecting relationships between query genes involved in retinitis pigmentosa etiopathogenesis. Edge colors: Co-expression (light purple), physical interaction (antique pink), genetic interaction (green), shared protein domains (golden yellow), pathway (light blue), predicted (orange) and common function (grey).

Fig. (2). Five genes extracted from the whole intersection of fifty causative ones, with the common feature of involvement in phototransduction. This figure shows the most relevant pathway common to five clustered genes, highlighting the importance of phototransduction function.

Fig. (3). BinGO particular of five genes in exam. Graph evidences the close relationship between them and physiological activities of vision ones (most of which included into fifty causatives of retinitis pigmentosa, in comparison with Fig. 3).

RCVRN: this gene encodes a member of the recoverin family of neuronal calcium sensors, which contains three calcium-binding EF-hand domains. It may delay phototransduction cascade termination in the retina by blocking the phosphorylation of photo-activated rhodopsin [31, 32]. Recoverin may be the antigen responsible for cancer-associated retinopathy [33].

GNB1 and GNGT1: heterotrimeric guanine nucleotide-binding proteins (G proteins), which transduce extracellular signals received by transmembrane receptors to effector proteins [34]. They are membrane bound GTPases that are linked to 7-TM receptors. Each G protein contains an α-, β- and γ-subunit and is bound to GDP in the “off” state. Ligand-receptor binding results in detachment of the G protein, switching it to an “on” state and permitting G_α activation of second messenger signalling cascades. Transducin is one of these proteins, found specifically in rod outer segments, where it mediates the activation of a cyclic GTP-specific guanosine monophosphate phosphodiesterase by rhodopsin [35]. GNB1 encodes a β subunit, while GNGT1 the gamma one [36, 37]. The α subunit is encoded by GNAT1 gene. The β and γ chains are required for the GTPase activity, for replacement of GDP by GTP, and for G protein-effector interaction (provided by RefSeq, Sep 2013).

GRK7: This gene encodes a member of the guanine nucleotide-binding protein (G protein)-coupled receptor kinase subfamily of the Ser/Thr protein kinase family (GRK) [38]. It is a retina-specific kinase involved in the shutoff of the photoresponse and adaptation to changing light conditions via cone opsin phosphorylation, including rhodopsin (RHO) [39]. It is believed that the movements of intracytoplasmic loop of activated RHO are responsible for the activation of the GRK7, in a way similar to GRK1 [40] (Fig. 5)).

Fig. (4). Phototransduction steps. 1) activation of the receptor protein in rods, that is rhodopsin (1 photon → 1 rhodopsin); 2) the activated receptor protein stimulates the G-protein transducin, with GTP converted to GDP in the process (1 rhodopsin → 100 transducins); 3) in turn, activated transducin activates the effector protein phosphodiesterase, converting cGMP to GMP (1 transducin → 100 PDE/s); 4) Falling concentrations of cGMP cause the transduction channels to close, decreasing Na+ current (1 PDE → 1000 GMP/s).

Fig. (5). Molecular mechanisms of GRK1 activation. GRK1 is a membrane – associated enzyme that has low affinity for Rho. After photoactivation, due to conformational changes in the receptor, GRK1 forms a complex with Rho* by associating with the secondo and third cytoplasmic loops of the receptor. According to one model, GRK1 becomes activated, dissociates, and phosphorilates the C - terminal region of many Rho. According to a second model, GRK1 remains bound and phosphorilates the nearby C – termini of Rho and Rho* at Ser334, Ser338 or Ser343.

ARRB1: Members of arrestin/beta-arrestin protein family are thought to participate in agonist-mediated desensitization of G-protein-coupled receptors and cause specific dampening of cellular responses to stimuli such as hormones, neurotransmitters, or sensory signals [41]. Arrestin beta 1 is a cytosolic protein and acts as a cofactor in the beta-adrenergic receptor kinase (BARK) mediated desensitization of beta-adrenergic receptors. During homologous desensitization, beta-arrestins bind to the GPRK-phosphorylated receptor and sterically preclude its coupling to the cognate G – protein (Fig. 6); the binding appears to require additional receptor determinants exposed only in the active receptor conformation [42]. The beta-arrestins target many receptors for internalization by acting as endocytic adapters (CLASPs, clathrin-associated sorting proteins) and recruiting the GPRCs to the adapter protein 2 complex 2 (AP-2) in clathrin-coated pits (CCPs). However, the extent of beta-arrestin involvement appears to vary significantly depending on the receptor, agonist and cell type. Internalized arrestin-receptor complexes traffic to intracellular endosomes, where they remain uncoupled from G-proteins [43]. Beta-arrestins function as multivalent adapter proteins that can act as signaling scaffold for MAPK pathways, like several other proteins as CCM proteins [44, 45], in the so-called beta-arrestin signalosomes [46]. Moreover, they can be involved in IGF1-stimulated AKT1 signaling leading to increased protection from apoptosis, in activation of the p38 MAPK signaling pathway and in actin bundle formation, in F2RL1-mediated cytoskeletal rearrangement and chemotaxis, and in AGTR1-mediated stress fiber formation by acting together with GNAQ to activate RHOA [47].