Dataset Information

Retinal Degeneration Caused by Rod-Specific Dhdds Ablation Occurs without Concomitant Inhibition of Protein N-Glycosylation.

ABSTRACT: Dehydrodolichyl diphosphate synthase (DHDDS) catalyzes the committed step in dolichol synthesis. Recessive mutations in DHDDS cause retinitis pigmentosa (RP59), resulting in blindness. We hypothesized that rod photoreceptor-specific ablation of Dhdds would cause retinal degeneration due to diminished dolichol-dependent protein N-glycosylation. Dhddsflx/flx mice were crossed with rod-specific Cre recombinase-expressing (Rho-iCre75) mice to generate rod-specific Dhdds knockout mice (Dhddsflx/flx iCre+). In vivo morphological and electrophysiological evaluation of Dhddsflx/flx iCre+ retinas revealed mild retinal dysfunction at postnatal (PN) 4 weeks, compared with age-matched controls; however, rapid photoreceptor degeneration ensued, resulting in almost complete loss of rods and cones by PN 6 weeks. Retina dolichol levels were markedly decreased by PN 4 weeks in Dhddsflx/flx iCre+ mice, relative to controls; despite this, N-glycosylation of retinal proteins, including opsin (the dominant rod-specific glycoprotein), persisted in Dhddsflx/flx iCre+ mice. These findings challenge the conventional mechanistic view of RP59 as a congenital disorder of glycosylation.

SUBMITTER: Ramachandra Rao S

PROVIDER: S-EPMC7287266 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Retinal Degeneration Caused by Rod-Specific Dhdds Ablation Occurs without Concomitant Inhibition of Protein N-Glycosylation.

Ramachandra Rao Sriganesh S Skelton Lara A LA Wu Fuguo F Onysk Agnieszka A Spolnik Grzegorz G Danikiewicz Witold W Butler Mark C MC Stacks Delores A DA Surmacz Liliana L Mu Xiuqian X Swiezewska Ewa E Pittler Steven J SJ Fliesler Steven J SJ

iScience 20200523 6

Dehydrodolichyl diphosphate synthase (DHDDS) catalyzes the committed step in dolichol synthesis. Recessive mutations in DHDDS cause retinitis pigmentosa (RP59), resulting in blindness. We hypothesized that rod photoreceptor-specific ablation of Dhdds would cause retinal degeneration due to diminished dolichol-dependent protein N-glycosylation. Dhdds<sup>flx/flx</sup> mice were crossed with rod-specific Cre recombinase-expressing (Rho-iCre75) mice to generate rod-specific Dhdds knockout mice (Dhd ...[more]

PMID: 32526701

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Project description:Dysfunction of Müller cells has been implicated in the pathogenesis of several retinal diseases. In order to understand the potential contribution of Müller cells to retinal disease better, we have developed a transgenic model in which foci of Müller cell ablation can be selectively induced. MicroRNAs (miRNAs), small non-coding RNAs that are involved in post-transcriptional modulation, have critical functions in various biological processes. The aim of this study was to profile differential expression of miRNAs and to examine changes in their target genes 2 weeks after Müller cell ablation. We identified 20 miRNAs using the miScript HC PCR array. Data analysis using two target gene prediction databases (TargetScan and mirTarBase) revealed 78 overlapping target genes. DAVID and KEGG pathway analysis suggested that the target genes were generally involved in cell apoptosis, p53, neurotrophin, calcium, chemokine and Jak-STAT signalling pathways. Changes in seven target genes including Cyclin D2, Caspase 9, insulin-like growth factor 1, IL-1 receptor-associated kinase (IRAK), calmodulin (CALM) and Janus kinase 2 (Jak2), were validated with qRT-PCR and western blots. The cellular localisation of cleaved-caspase 9, Cyclin D2, Jak2 and CALM was examined by immunofluorescence studies. We found that the transcription of some miRNAs was positively, rather than negatively, correlated with their target genes. After confirming that overexpressed miR-133a-3p was localised to the outer nuclear layer in the damaged retina, we validated the correlation between miR-133a-3p and one of its predicted target genes, cyclin D2, with a luciferase assay in 661 photoreceptor cells. Results revealed by miRNA profiling, target gene analysis and validation were generally consistent with our previous findings that selective Müller cell ablation causes photoreceptor degeneration and neuroinflammation. Our data on alterations of miRNAs and their target gene expression after Müller cell ablation provide further insights into the potential role of Müller cell dysfunction in retinal disease.

Dataset Information

Retinal Degeneration Caused by Rod-Specific Dhdds Ablation Occurs without Concomitant Inhibition of Protein N-Glycosylation.

Publications

Retinal Degeneration Caused by Rod-Specific Dhdds Ablation Occurs without Concomitant Inhibition of Protein N-Glycosylation.

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