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Biology subjects

McKie, L.

Publications and source records attributed to McKie, L..

6 recordsLinked to original sources

Autophagy disruption and mitochondrial stress precede photoreceptor necroptosis in multiple mouse models of inherited retinal disorders.

Inherited retinal diseases (IRDs) are a leading cause of blindness worldwide. One of the greatest barriers to developing treatments for IRDs is the heterogeneity of these disorders, with causative mutations identified in over 280 genes. It is therefore a priority to find therapies applicable to a broad range of genetic causes. To do so requires a greater understanding of the common or overlapping molecular pathways that lead to photoreceptor death in IRDs and the molecular processes through which they converge. Here, we characterise the contribution of different cell death mechanisms to photoreceptor degeneration and loss throughout disease progression in humanised mouse models of IRDs. Using single-cell transcriptomics, we identify common transcriptional signatures in degenerating photoreceptors. Further, we show that in genetically and functionally distinct IRD models, common early defects in autophagy and mitochondrial damage exist, triggering photoreceptor cell death by necroptosis in later disease stages. These results suggest that, regardless of the underlying genetic cause, these pathways likely contribute to cell death in IRDs. These insights provide potential therapeutic targets for novel, gene-agnostic treatments for IRDs applicable to the majority of patients.

cell biology↗

Ciliary tip actin dynamics regulate the cadenceof photoreceptor disc formation

As signalling organelles, primary cilia regulate their membrane G protein-coupled receptor (GPCR) content by ectocytosis, a process requiring localised actin dynamics at their tip to alter membrane shape.(1, 2) Mammalian photoreceptor outer segments comprise an expanse of folded membranes (discs) at the tip of highly-specialised connecting cilia (CC), in which photosensitive GPCRs like rhodopsin are concentrated. In an extraordinary feat of biology, outer segment discs are shed and remade daily.(3) Defects in this process, due to genetic mutations, cause retinitis pigmentosa (RP), an untreatable, blinding disease. The mechanism by which photoreceptor cilia generate outer segments is therefore fundamental for vision yet poorly understood. Here, we show the membrane deformation required for outer segment disc genesis is driven by dynamic changes in the actin cytoskeleton in a process akin to ectocytosis. Further, we show RPGR, a leading causal RP gene, regulates activity of actin binding proteins crucial to this process. Disc genesis is compromised in Rpgr mouse models, slowing the actin dynamics required for timely disc formation, leading to aborted membrane shedding as ectosome-like vesicles, photoreceptor death and visual loss. Manipulation of actin dynamics partially rescues the phenotype, suggesting this pathway could be targeted therapeutically. These findings help define how actin-mediated dynamics control outer segment turnover.

cell biology↗

The Dct-/- mouse model to unravel retinogenesis misregulation in patients with albinism

We have recently identified DCT encoding dopachrome tautomerase (DCT) as the 8th gene for oculocutaneous albinism (OCA). Patients with loss of function of DCT suffer from eye hypopigmentation and retinal dystrophy. Here we investigate the eye phenotype in Dct-/- mice. We show that their retinal pigmented epithelium (RPE) is severely hypopigmented from early stages contrasting with the darker melanocytic tissues. Multimodal imaging reveals specific RPE cellular defects. Melanosomes are fewer with correct subcellular localization but disrupted melanisation. RPE cell size is globally increased and heterogeneous. P-cadherin labeling of Dct-/- newborn RPE reveals a defect in adherens junctions similar to what has been described in tyrosinase-deficient Tyrc/c embryos. The first intermediate of melanin biosynthesis, dihydroxyphenylalanine (L-Dopa), which is thought to control retinogenesis, is detected in substantial yet significantly reduced amounts in Dct-/- postnatal mouse eyecups. L-Dopa synthesis in the RPE alone remains to be evaluated during the critical period of retinogenesis. The Dct-/- mouse should prove useful in understanding the molecular regulation of retinal development and aging of the hypopigmented eye. This may guide therapeutic strategies to prevent vision deficits in patients with albinism.

genetics↗

Centriolar satellites expedite mother centriole remodeling to promote ciliogenesis.

Centrosomes are orbited by centriolar satellites, dynamic multiprotein assemblies nucleated by PCM1. To study the requirement for centriolar satellites, we generated mice lacking PCM1. Pcm1-/- mice display partially penetrant perinatal lethality with survivors exhibiting hydrocephalus, oligospermia and cerebellar hypoplasia, as well as variable expressivity of other ciliopathy features including cystic kidneys. Pcm1-/- multiciliated ependymal cells and PCM1-/- retinal pigmented epithelial 1 (RPE1) cells showed reduced ciliogenesis. PCM1-/- RPE1 cells displayed reduced docking of the mother centriole to the ciliary vesicle and removal of CP110 and CEP97 from the distal mother centriole, indicating compromized early ciliogenesis. We show these molecular cascades are maintained in vivo, and we suggest that the cellular threshold to trigger ciliogenesis varies between cell types. We propose that PCM1 and centriolar satellites facilitate efficient trafficking of proteins to and from centrioles, inducing the departure of CP110 and CEP97 to initiate ciliogenesis.

cell biology↗

A Mouse Model of Brittle Cornea Syndrome caused by mutation in Zfp469.

Brittle Cornea Syndrome (BCS) is a rare recessive condition characterised by extreme thinning of the cornea and sclera. BCS results from loss-of-function mutations in the poorly understood genes ZNF469 or PRDM5. In order to determine the function of ZNF469 and to elucidate pathogenic mechanisms, we used genome editing to recapitulate a human ZNF469 BCS mutation in the orthologous mouse gene, Zfp469. Ophthalmic phenotyping showed that homozygous Zfp469 mutation causes significant central and peripheral corneal thinning arising from reduced stromal thickness. Expression of key components of the corneal stroma in primary keratocytes from Zfp469BCS/BCS mice is affected, including decreased Col1a1 and Col1a2 expression. This alters the type I:type V collagen ratio and results in collagen fibrils with smaller diameter and increased fibril density in homozygous mutant corneas, correlating with decreased biomechanical strength in the cornea. Cell-derived matrices generated by primary keratocytes show reduced deposition of type I collagen offering an in vitro model for stromal dysfunction. Work remains to determine whether modulating ZNF469 activity will have therapeutic benefit in BCS or in conditions such as keratoconus where the cornea thins progressively. Summary statementA mouse model of Brittle Cornea Syndrome was created to elucidate molecular mechanisms underlying pathology of this rare connective tissue disorder in which extremely thin corneas rupture, causing irreversible blindness.

genetics↗

Dopachrome tautomerase variants in patients with oculocutaneous albinism.

PurposeAlbinism is a clinically and genetically heterogeneous condition. Despite analysis of the nineteen known genes, [~]30% patients remain unsolved. We aimed to identify new genes involved in albinism. MethodsWe sequenced a panel of genes with known or predicted involvement in melanogenesis in 230 unsolved albinism patients. ResultsWe identified variants in the Dopachrome tautomerase (DCT) gene in two patients. One was compound heterozygous for a 14 bp deletion in exon 9 and c.118T>A p.(Cys40Ser). The second was homozygous for c.183C>G p.(Cys61Trp). Both patients had mild hair and skin hypopigmentation, and classical ocular features. CRISPR/Cas9 was used in C57BL/6J mice to create mutations identical to the missense mutations carried by the patients, along with one loss-of-function indel mutation. When bred to homozygosity the three mutations revealed hypopigmentation of the coat, milder for Cys40Ser compared to Cys61Trp or the frameshift mutation. Histological analysis identified significant hypopigmentation of the retinal pigmented epithelium (RPE) indicating that defective RPE melanogenesis could be associated with eye and vision defects. DCT loss of function in zebrafish embryos elicited hypopigmentation both in melanocytes and RPE cells. ConclusionsDCT is the gene for a new type of oculocutaneous albinism that we propose to name OCA8.

genetics↗