15.6 Perspective

receptors in the RPE is not merely a consequence of ligand-induced stabilization or of anchorage to the MerTK dependent engulfment machinery. Rather, we propose that RPE cells use a trafficking pathway to specifically sort αvβ5 to the apical surface. This pathway is likely to recognize motifs of the β5 cytoplasmic domain. Comparing trafficking and complex formation among αvβ5 receptors with cytoplasmic deletions and point mutations will be our future approach to identify trafficking-relevant residues and motifs of the β5 integrin cytoplasmic tail.

Acknowledgment This work was supported by NIH grant R01-EY13295 to SCF.

References

Calderwood DA (2004) Talin controls integrin activation. Biochem Soc Trans 32(Pt3):434–437 Cook B, Lewis GP et al (1995) Apoptotic photoreceptor degeneration in experimental retinal

detachment. Invest Ophthalmol Vis Sci 36(6):990–996

Deora AA, Gravotta D et al (2004) The basolateral targeting signal of CD147 (EMMPRIN) consists of a single leucine and is not recognized by retinal pigment epithelium. Mol Biol Cell 15(9):4148–4165

Eliceiri BP, Puente XS et al (2002) Src-mediated coupling of focal adhesion kinase to integrin alpha(v)beta5 in vascular endothelial growth factor signaling. J Cell Biol 157(1): 149–160

Finnemann SC (2003) Focal adhesion kinase signaling promotes phagocytosis of integrin-bound photoreceptors. EMBO J 22(16):4143–4154

Finnemann SC, Bonilha VL et al (1997) Phagocytosis of rod outer segments by retinal pigment epithelial cells requires alpha(v)beta5 integrin for binding but not for internalization. Proc Natl Acad Sci USA 94(24):12932–12937

Finnemann SC, Chang Y (2008) Photoreceptor-RPE interactions physiology and molecular mechanisms. In: Tombran-Tink J, Barnstable CJ (eds) From ophthalmology research: visual transduction and non-visual light perception. Humana Press, Totowa, NJ

Fisher SK, Lewis GP et al (2005) Cellular remodeling in mammalian retina: results from studies of experimental retinal detachment. Prog Retin Eye Res 24(3):395–431

Frisch SM, Screaton RA (2001) Anoikis mechanisms. Curr Opin Cell Biol 13(5):555–562 Gundersen D, Powell SK et al (1993) Apical polarization of N-CAM in retinal pigment epithelium

is dependent on contact with the neural retina. J Cell Biol 121(2):335–343

Horwitz A, Duggan K et al (1986) Interaction of plasma membrane fibronectin receptor with talin – a transmembrane linkage. Nature 320(6062):531–533

Hunziker W, Fumey C (1994) A di-leucine motif mediates endocytosis and basolateral sorting of macrophage IgG Fc receptors in MDCK cells. EMBO J 13(13):2963–2969

Legate KR, Fassler R (2009) Mechanisms that regulate adaptor binding to beta-integrin cytoplasmic tails. J Cell Sci 122(Pt 2):187–198

Marmorstein AD, Gan YC et al (1998) Apical polarity of N-CAM and EMMPRIN in retinal pigment epithelium resulting from suppression of basolateral signal recognition. J Cell Biol 142(3):697–710

Nandrot EF, Anand M et al (2006) Novel role for alphavbeta5-integrin in retinal adhesion and its diurnal peak. Am J Physiol Cell Physiol 290(4):C1256–C1262

Nandrot EF, Anand M et al (2007) Essential role for MFG-E8 as ligand for alphavbeta5 integrin in diurnal retinal phagocytosis. Proc Natl Acad Sci USA 104(29):12005–12010

Nandrot EF, Kim Y et al (2004) Loss of synchronized retinal phagocytosis and age-related blindness in mice lacking alphavbeta5 integrin. J Exp Med 200(12):1539–1545

Singh S, D’Mello V et al (2007) A NPxY-independent beta5 integrin activation signal regulates phagocytosis of apoptotic cells. Biochem Biophys Res Commun 364(3):540–548

Chapter 16

Mertk in Daily Retinal Phagocytosis: A History

in the Making

Emeline F. Nandrot and Eric M. Dufour

Abstract It took 62 years from the description of the retinal dystrophy in rats from the Royal College of Surgeons (RCS) strain to the discovery of the molecular defect underlying the phenotype. Phagocytosis of photoreceptor outer segments (POS) by retinal pigment epithelial (RPE) cells follows a daily rhythm with a peak of activity 1.5–2 h after light onset for rod photoreceptors. We identified a deletion in the Mer tyrosine kinase (MerTK) receptor in RCS rat that abolishes internalization of POS by RPE cells. Accumulation of debris in the subretinal space then leads to drastic photoreceptor degeneration and rapid loss of vision. Interestingly, in wild-type mice and rats, MerTK is phosphorylated at the time of the phagocytic peak. We also demonstrated that the couple αvβ5 integrin receptor and MFG-E8 ligand synchronizes daily retinal phagocytosis. Indeed, when either one is absent in knockout mice, phagocytosis follows steady-state levels, and peak activation of integrin-associated protein and of MerTK does not occur. We now have a more precise picture of the sequence of molecular events governing retinal phagocytosis. However, requirement of MerTK ligands in vivo and linked signaling pathways still remain elusive so far.

16.1 Introduction

The retina is constituted of post-mitotic cells organized in layers that differentiate during eye development. Among these, two adjacent cell types of the outer retina, photoreceptors (PR) and retinal pigment epithelial (RPE) cells, interact with each other functionally and are highly inter-dependant. RPE cells form a highly specialized monolayer endorsing many activities to ensure function and integrity of the neural retina and therefore vision (review: Strauss 2005). On their apical side, RPE cells extend microvilli that ensheath the outer segment of rod and cone PRs.

E.F. Nandrot (B)

Centre de Recherche Institut de la Vision, UMR_S968, Inserm, UPMC University of Paris 06, 17 rue Moreau, 75012 Paris, France

e-mail: emeline.nandrot@inserm.fr

Medicine and Biology 664, DOI 10.1007/978-1-4419-1399-9_16,C Springer Science+Business Media, LLC 2010

Photoreceptor outer segment (POS) structure consists of stacked membranous disks containing the phototransduction machinery. POS are subjected to sustained light exposure thus generating high levels of oxidative stress. In order to ensure lifelong function, both photoreceptor rods and cones continuously replace their outer segments (Young 1967). New membranous disks are initiated in the connecting cilium, inserted at the base of the inner segment, and progressively move towards the distal end of the POS. As a counterpart regulatory mechanism, PRs shed their most distal tips every day to keep constant cell length. The daily shedding of POS tips is controlled by strong circadian rhythms influenced by the dark-light cycle (Goldman et al.1980). Animal studies in rod-dominant species revealed that rods mainly shed their outer segments within 2 h after the light onset (LaVail and Mullen 1976). Upon rhythmic shedding, RPE cells respond with a burst of synchronized phagocytosis that efficiently digests POS and recycles their components (Young and Bok 1969). In mammals, each RPE cell serves approximately 30 POS that shed 7% of their outer segment mass every day (Young 1967), thus rendering RPE cells the most active phagocytes in the body.

16.2 RCS Rat and MerTK Receptor: An Intimate Story

Over 70 years ago, Margherita C. Bourne described a retinal degeneration occurring in rats affected by cataracts (Bourne et al. 1938) (Fig. 16.1a). She started breeding affected rats together, and the following year, she showed that the defect was transmitted as an autosomic recessive trait (Bourne and Grüneberg 1939). This animal model would be known later as the Royal College of Surgeons (RCS) rat strain.

The first defect detected is the accumulation of POS disk-like structures observed at 12 days on electron micrographs and 18 days with light microscopy (Dowling and Sidman 1962). Electroretinogram (ERG) retinal responses begin to decrease at 18 days until no more response can be detected at 3 months (Dowling and Sidman 1962; Nandrot et al. 2000) (Fig. 16.1b). PR cell death starts around 3 weeks of age until the complete loss of PR cells at 3 months (Bourne et al. 1938; Dowling and Sidman 1962; Nandrot et al. 2000) (Fig. 16.1a). Then, further disruption of the whole retina structure and cell degeneration spreads to all the other cell layers.

Mullen and LaVail (1976) used chimeras of non-pigmented RCS and pigmented control rats and showed that RPE cells completely fail to engulf POS, which in turn causes the drastic accumulation of POS debris in the subretinal space. Consistently, Edwards and Szamier (1977) demonstrated in vitro that the RPE cells defect is POS-specific, RPE cells being able to phagocytose latex beads. The exact phagocytosis disruption was pinpointed by Chaitin and Hall (1983) and further confirmed by Hall and Abrams (1987): RCS RPE cells bound POS properly but were unable to subsequently internalize them.

It took almost 15 more years to identify the molecular defect underlying the RCS rat retinal dystrophy. The gene was first localized to chromosome 3 by MM LaVail (1981) using co-transmission of enzymatic markers with the retinal dystrophy. In