Disruption of the retinitis pigmentosa 28 gene Fam161a in mice affects photoreceptor ciliary structure and leads to progressive retinal degeneration

Abstract

Mutations in the FAM161A gene were previously identified as the cause for autosomal-recessive retinitis pigmentosa 28. To study the effects of Fam161a dysfunction in vivo, we generated gene-trapped Fam161a(GT/GT) mice with a disruption of its C-terminal domain essential for protein-protein interactions. We confirmed the absence of the full-length Fam161a protein in the retina of Fam161a(GT/GT) mice using western blots and showed weak expression of a truncated Fam161a protein by immunohistochemistry. Histological analyses demonstrated that photoreceptor segments were disorganized in young Fam161a(GT/GT) mice and that the outer retina was completely lost at 6 months of age. Reactive microglia appeared in the outer retina and electroretinography showed an early loss of photoreceptor function in 4-month-old Fam161a(GT/GT) animals. Light and electron microscopy revealed a remarkable phenotype of a significantly shortened connecting cilium, spread ciliary microtubule doublets and disturbed disk organization in Fam161a(GT/GT) photoreceptor cells. Co-immunolabeling experiments demonstrated reduced expression and mislocalization of centrin 3 and disturbed targeting of the Fam161a interactors lebercilin and Cep290, which were restricted to the basal body and proximal connecting cilium in Fam161a(GT/GT) retinas. Moreover, we identified misrouting of the outer segment cargo proteins opsin and rds/peripherin 2 in Fam161a(GT/GT) mice. In conclusion, our results suggest a critical role for the C-terminal domain of Fam161a for molecular interactions and integrity of the connecting cilium. Fam161a is required for the molecular delivery into the outer segment cilium, a function which is essential for outer segment disk formation and ultimately visual function.

Alterations in the balance of tubulin glycylation and glutamylation in photoreceptors leads to retinal degeneration

“…Our study shows that mutation of a single glycylase gene, Ttll3, is sufficient to induce slow retinal degeneration in mice, as revealed by shortened connecting cilia (Karlstetter et al, 2014), a massive activation of Müller glia, indicative of retinal stress (Honjo et al, 2000;Iandiev et al, 2006), and a remarkable increase of photoreceptor apoptosis. The degeneration could be related to a partial dysfunction of the IFT in the connecting cilia, which is considered a possible cause of retinal degeneration (Alfinito and Townes-Anderson, 2002;Boubakri et al, 2016;Bujakowska et al, 2015;Hollingsworth and Gross, 2012;Jiang et al, 2009).…”

“…As shortening of connecting cilia could be a sign of retinal degeneration (Karlstetter et al, 2014), we analysed retinas of Ttll3 −/− mice for signs of degeneration. First, we determined the distribution of rhodopsin in retina sections of 12-month-old mice.…”

Comprehensive analysis of mouse retinal mononuclear phagocytes

“…We have previously characterized the macrophage/microglia WAP domain protein (AMWAP) as a regulator of pro-inflammatory response 37,38 . AMWAP transcription is rapidly induced in microglia from different retinal disease mouse models, including inherited photoreceptor dystrophies and different light challenge paradigms 7,32,33,39 . Another good marker to detect microglia reactivity is CCchemokine ligand 2 (CCL2, alias monocyte chemoattractant protein 1, MCP-1), which has a key role in pro-inflammatory mononuclear phagocyte chemotaxis and migration in the retina 14,40 .…”

Diverse clinical phenotypes associated with a nonsense mutation in FAM161A

“…4 Although it has been shown that FAM161A is required for routing of outer segment proteins, such as opsin and rds/peripherin 2, its precise function remains unclear. 6,7 We aimed to gain insights into the frequency of FAM161A mutations in patients with RP seen at a single UK centre and to characterise the retinal phenotype in individuals with FAM161A-related disease.…”