CEP290 myosin-tail homology domain is essential for protein confinement between inner and outer segments in photoreceptors

Abstract

Mutations in CEP290 cause various ciliopathies involving retinal degeneration. CEP290 proteins localize to the ciliary transition zone and are thought to act as a gatekeeper that controls ciliary protein trafficking. However, precise roles of CEP290 in photoreceptors and pathomechanisms of retinal degeneration in CEP290-associated ciliopathies are not sufficiently understood. Using Cep290 conditional mutant mice, in which the C-terminal myosin-tail homology domain is disrupted after the connecting cilium is assembled, we show that CEP290, more specifically the myosin-tail homology domain of CEP290, is essential for protein confinement between the inner and the outer segments.Inner segment plasma membrane proteins including STX3, SNAP25, and IMPG2 rapidly accumulate in the outer segment upon disruption of the myosin-tail homology domain. In contrast, localization of endomembrane proteins is not altered. Trafficking and confinement of most outer segment-resident proteins appear to be unaffected or only minimally affected in this mouse model. One notable exception is RHO, which exhibits severe mislocalization to inner segments from the initial stage of degeneration.Similar mislocalization phenotypes were observed in rd16 mice. These results suggest that failure of protein confinement at the connecting cilium and consequent accumulation of inner segment membrane proteins in the outer segment combined with insufficient RHO delivery is part of the disease mechanisms that cause retinal degeneration in CEP290-associated ciliopathies. Our study provides insights into the pathomechanisms of retinal degenerations associated with compromised ciliary gates.the absence of CEP290 in these organisms, ciliary protein compositions are altered [4,6]. In mammalian primary cilia, CEP290 is localized to the transition zone [7][8][9], and loss of CEP290 reduces ARL13B and ADCY3 levels within cilia while increasing the rate of ciliary entry of SMO in fibroblasts [10]. These studies establish the current model for CEP290 function: a ciliary gatekeeper that regulates protein trafficking in and out of the ciliary compartment at the transition zone [4][5][6][7][10][11][12][13]. In photoreceptors, CEP290 is localized to the connecting cilium [14,15] and expected to control protein movement between the inner and the outer segments.Mutations in human CEP290 cause various ciliopathies ranging from isolated retinal dystrophy (e.g.Leber congenital amaurosis (LCA)) to syndromic diseases such as neonatal lethal Meckel-Gruber Syndrome (MKS) with multi-organ malformations [16][17][18][19][20][21][22]. Despite considerable variations in phenotypic severity, retinopathy is present in almost all cases regardless of the involvement of other organs. This suggests that photoreceptors are particularly susceptible to deficiencies in CEP290 function. Based on the ciliary gatekeeper model, anticipated functions of CEP290 in photoreceptors include i) permitting or facilitating entry of outer segment-bound proteins into the outer segment, ii) blocking unauthoriz...

Super-resolution microscopy reveals photoreceptor-specific subciliary location and function of ciliopathy-associated protein CEP290

“…Joubert syndrome is a syndromic ciliopathy characterized by nephronophthisis, cerebellar vermis aplasia, and retinal degeneration (58). These Cep290 tm1.1Jgg/tm1.1Jgg g animals will be referred to throughout the rest of the paper as a near-null (NN), because an alternatively spliced variant of the mutant allele may result in low residual levels of a truncated CEP290 (59).…”

Superresolution microscopy reveals photoreceptor-specific subciliary location and function of Cep290

“…The Cep290 tm1.1Jgg animals present with rapid photoreceptor degeneration and vermal hypoplasia (54). The Cep290 tm1.1Jgg allele is described as a null allele; however, an alternatively spliced variant of the mutant allele may result in low residual levels of a truncated CEP290 (56). Immunostaining of retinal sections revealed little or no labeling with the CEP290 antibody in Cep290 tm1.1Jgg retinas at P10 ( Figure 8A-B).…”