Single-cell transcriptomic atlas of the human retina identifies cell types associated with age-related macular degeneration

Menon, Madhvi; Mohammadi, Shahin; Dávila-Velderrain, José; Goods, Brittany A.; Cadwell, Tanina D.; Yu, Xiuping; Stemmer‐Rachamimov, Anat; Shalek, Alex K.; Love, J. Christopher; Kellis, M; Hafler, Brian P.

doi:10.1038/s41467-019-12780-8

Cited by 248 publications

(253 citation statements)

References 42 publications

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“…The loss of NMNAT2 induces an increase in NMN that is hypothesized to activate SARM1-dependent axon degeneration 53,54 . Our metabolomic analysis revealed that NMN is increased in the NMNAT1-deficient retinas ( Figure 1D), and previous studies have detected SARM1 in mouse and bovine photoreceptor cells [55][56][57] . These results raise the possibility that the increased retinal NMN activates SARM1 and induces NAD + loss and cellular degeneration in the retina.…”

Section: Resultssupporting

confidence: 86%

“…Finally, we defined the molecular mechanism by which NMNAT1 promotes photoreceptor function and survival. In Single-cell transcriptomic RNA analysis also found Nmnat1 in rods and cones 57,61 . Consistent with an extranuclear role for NMNAT1 in photoreceptors in these studies, cytosolic NMNAT2 was either not identified or was found at much lower levels than NMNAT1.…”

Section: Discussionmentioning

confidence: 84%

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SARM1 depletion rescues NMNAT1 dependent photoreceptor cell death and retinal degeneration

Sasaki

Kakita

Kubota

et al. 2020

Preprint

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Leber congenital amaurosis type 9 (LCA9) is an autosomal recessive, early onset retinal neurodegenerative disease caused by mutations in the gene encoding the nuclear NAD + synthesis enzyme NMNAT1. Despite the ubiquitous expression of NMNAT1 and its role in NAD + homeostasis, LCA9 patients do not manifest pathologies other than retinal degeneration.To investigate the mechanism of degeneration, we examined retinas of developing and adult mice with conditional or tissue-specific NMNAT1 loss. Widespread NMNAT1 depletion in adult mice resulted in loss of photoreceptors, indicating these cells are exquisitely vulnerable to NMNAT1 loss. NMNAT1 is required within the photoreceptor, as conditional deletion of NMNAT1 in photoreceptors but not retinal pigment epithelial cells is sufficient to cause photoreceptor neurodegeneration and vision loss. Moreover, delivery of NMNAT1 into eyes of adult mice lacking NMNAT1 using a modified AAV8 vector containing a photoreceptor-specific promoter rescued the retinal degeneration phenotype and partially restored vision. Finally, we defined the molecular mechanism driving photoreceptor cell death. Loss of NMNAT1 activates SARM1, an inducible NADase best known as the central executioner of axon degeneration. SARM1 is required for the photoreceptor death and vision loss that occurs following NMNAT1 deletion. This surprising finding demonstrates that the essential function of NMNAT1 in photoreceptors is to inhibit SARM1, and establishes a commonality of mechanism between axonal degeneration and photoreceptor neurodegeneration. These results define a novel SARM1-dependent photoreceptor cell death pathway that is active in the setting of dysregulated NAD + metabolism and identifies SARM1 as a therapeutic candidate for the treatment of retinal degeneration.

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Section: Resultssupporting

confidence: 86%

Section: Discussionmentioning

confidence: 84%

SARM1 depletion rescues NMNAT1 dependent photoreceptor cell death and retinal degeneration

Sasaki

Kakita

Kubota

et al. 2020

Preprint

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“…Single-cell RNA sequencing (scRNA-seq) is a powerful tool to study development, tissue, and disease biology [1][2][3][4][5][6][7][8][9] . scRNA-seq analyses currently rely on the availability of high quality genome annotations to define cell features and to perform cell clustering, dimensionality reduction, differential gene expression, and other analyses [10][11][12][13][14] . Identifying genes and correctly annotating their locations and coding regions within the genome is a difficult task that collaborative projects such as Ensembl, ENCODE, and GENCODE are striving to achieve.…”

Section: Introductionmentioning

confidence: 99%

Uncovering Transcriptional Dark Matter via Gene Annotation Independent Single-Cell RNA Sequencing Analysis

Wang

Mantri

Chou

et al. 2020

Preprint

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Single-cell RNA sequencing (scRNA-seq) enables the study of cell biology with high resolution. scRNA-seq expression analyses rely on the availability of a high quality annotation of genes in the genome. Yet, as we show here with scRNA-seq experiments and analyses spanning human, mouse, chicken, mole rat, lemur and sea urchin, gene annotations often fail to cover the full transcriptome of every cell type at every stage of development, in particular for organisms that are not routinely studied. To overcome this hurdle, we created a scRNA-seq analysis routine that recovers biologically relevant information beyond the scope of the best available gene annotation. This is achieved by performing single-cell expression analysis on any region in the genome for which transcriptional products are detected. Our routine identifies transcriptionally active regions (TARs) using a hidden Markov model, generates a matrix of expression levels for all TARs across all cells in a dataset, performs single-cell TAR expression analysis to identify TARs that are biologically significant, and then annotates biologically significant TARs using gene homology analysis. This procedure leverages single-cell expression analyses as a filter to direct annotation efforts to biologically significant transcripts in complex tissues and thereby uncovers biology to which scRNA-seq would otherwise be in the dark.

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“…As an extension of the CNS, the retina is a commonly used model to study different neuron types [16]. The retina is composed of several neuron types that are well characterized from early development through adult stages [17][18][19][20][21][22][23][24][25][26][27]. Within the retina, the RGCs primarily relay visual information to the brain [28].…”

Section: Introductionmentioning

confidence: 99%

Optimized culture of retinal ganglion cells and amacrine cells from adult mice

Park

Snook

Zhuang

et al. 2020

Preprint

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Cell culture is widely utilized to study the cellular and molecular biology of different neuronal cell populations. Current techniques to study enriched neurons in vitro are primarily limited to embryonic/neonatal animals and induced pluripotent stem cells (iPSC). Although the use of these cultures is valuable, the accessibility of purified primary adult neuronal cultures would allow for improved assessment of certain neurological diseases and pathways at the cellular level. Using a modified 7-step immunopanning technique to isolate for retinal ganglion cells (RGCs) and amacrine cells (ACs) from adult mouse retinas, we have successfully developed a model of neuronal culture that maintains for at least one week. Isolations of Thy1.2 + cells are enriched for RGCs, with the isolation cell yield being congruent to the theoretical yield of RGCs in a mouse retina. ACs of two different populations (CD15 + and CD57 + ) can also be isolated. The populations of these three adult neurons in culture are healthy, with neurite outgrowths in some cases greater than 500µm in length. Optimization of culture conditions for RGCs and CD15 + cells revealed that neuronal survival and the likelihood of neurite outgrowth respond inversely to different culture media. Serially diluted concentrations of puromycin decreased cultured adult RGCs in a dose-dependent manner, demonstrating the potential usefulness of these adult neuronal cultures in screening assays. This novel culture system can be used to model adult neurons in vivo. Studies can now be expanded in conjunction with other methodologies to study the neurobiology of function, aging, and diseases.

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Single-cell transcriptomic atlas of the human retina identifies cell types associated with age-related macular degeneration

Cited by 248 publications

References 42 publications

SARM1 depletion rescues NMNAT1 dependent photoreceptor cell death and retinal degeneration

SARM1 depletion rescues NMNAT1 dependent photoreceptor cell death and retinal degeneration

Uncovering Transcriptional Dark Matter via Gene Annotation Independent Single-Cell RNA Sequencing Analysis

Optimized culture of retinal ganglion cells and amacrine cells from adult mice

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