Isolation and Comparative Transcriptome Analysis of Human Fetal and iPSC-Derived Cone Photoreceptor Cells

Welby, Emily; Lakowski, Jörn; Foggia, Valentina Di; Budinger, Dimitri; Gonzalez‐Cordero, Anai; Lun, Aaron T. L.; Epstein, Michael P.; Patel, Aara; Cuevas, Elisa; Kruczek, Kamil; Naeem, Aabgeena; Minneci, Federico; Hubank, Michael; Jones, David T.; Marioni, John C.; Ali, Robin R.; Sowden, Jane C.

doi:10.1016/j.stemcr.2017.10.018

Cited by 96 publications

(124 citation statements)

References 51 publications

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“…Correlation matrix to benchmark hiPSC‐derived cone photoreceptors (week 15 and week 20; Welby et al , ), fetal cone photoreceptors (Welby et al , ), adult retina (Phillips et al , ), and the human Müller glia cell line MIO‐M1 against all retinal cell types identified in this human neural retina atlas.…”

Section: Resultsmentioning

confidence: 99%

“… Correlation analysis of scRNA‐seq data of hiPSC‐derived cone photoreceptors (week 15) against fetal cone photoreceptors (Welby et al , ), as well as adult cone and rod photoreceptors from this human neural retina atlas. Principal component analysis to assess transcriptome similarity of hiPSC‐derived cone photoreceptors to fetal and adult photoreceptors. t‐SNE analysis of the human Müller glia cell line MIO‐M1 with the retinal cell types identified in this human neural retina atlas. Correlation analysis of MIO‐M1 with all major human retinal cell types. Top ranked differentially expressed genes identified in MIO‐M1 compared to other retinal cell types based on logistic regression score. …”

Section: Resultsmentioning

confidence: 99%

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A single‐cell transcriptome atlas of the adult human retina

et al. 2019

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The retina is a specialized neural tissue that senses light and initiates image processing. Although the functional organization of specific retina cells has been well studied, the molecular profile of many cell types remains unclear in humans. To comprehensively profile the human retina, we performed single‐cell RNA sequencing on 20,009 cells from three donors and compiled a reference transcriptome atlas. Using unsupervised clustering analysis, we identified 18 transcriptionally distinct cell populations representing all known neural retinal cells: rod photoreceptors, cone photoreceptors, Müller glia, bipolar cells, amacrine cells, retinal ganglion cells, horizontal cells, astrocytes, and microglia. Our data captured molecular profiles for healthy and putative early degenerating rod photoreceptors, and revealed the loss of MALAT1 expression with longer post‐mortem time, which potentially suggested a novel role of MALAT1 in rod photoreceptor degeneration. We have demonstrated the use of this retina transcriptome atlas to benchmark pluripotent stem cell‐derived cone photoreceptors and an adult Müller glia cell line. This work provides an important reference with unprecedented insights into the transcriptional landscape of human retinal cells, which is fundamental to understanding retinal biology and disease.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

A single‐cell transcriptome atlas of the adult human retina

et al. 2019

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“…For the comparison of retinal datasets, differential expression output data was obtained from each report (Kim et al, 2016;Mo et al, 2016;Welby et al). The data was cross-referenced to the genes obtained in the analysis from this paper and are detailed in Supp.…”

Section: Data Comparisonsmentioning

confidence: 99%

Identification of genes with enriched expression in early developing mouse cone photoreceptors

Buenaventura

Corseri

Emerson

2019

Preprint

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26Cone photoreceptors are the critical first cells that mediate high acuity vision. Despite 27 their importance and their potential use in cell-based therapies for retinal diseases, 28 there is a lack of knowledge about the early developmental stages of these cells. Here 29 we characterize the expression of the homeobox transcription factor Lhx4 as an early 30 and enriched cone photoreceptor expressed gene in both chicken and mouse. A Lhx4 31 GFP reporter mouse was found to recapitulate this early cone photoreceptor expression 32 and was used to purify and profile embryonic mouse cone photoreceptors by single cell 33 RNA sequencing. This enrichment in cone photoreceptors allowed for the robust 34 identification of genes associated with the early cone transcriptome and also identified 35 subpopulations of these cells. A comparison to previously reported datasets allowed the 36 classification of genes according to developmental timing, cell type specificity, and 37 whether they were regulated by the rod transcription factor Nrl. This analysis has 38 extended the set of known early cone enriched genes and identified those that are 39 regulated independently of Nrl. This report furthers our knowledge of the transcriptional 40 events that occur in early cone photoreceptors. 41 42 43 44 86 are not completely transformed into cones.87 88 89 RESULTS 90 LHX4 is present in early cone photoreceptors in the chicken retina 91 Recently, we established the transcriptional profile of retinal progenitor cells 92 (RPCs), defined by the activity of the ThrbCRM1 element, that are biased towards the 93 cone and horizontal cell (HC) fate in the early chick retina 3 . Using this dataset, we 94 screened for potential cone-enriched transcripts that could serve as markers for early 95 cones. After establishing a criterion for >1.5-fold change score between cone/HC RPCs 96 and other concurrent populations (enriched in "Other early retinal progenitors") we 97 selected for transcription factors (TFs) enriched in the cone/HC RPCs (Fig 1 A). We 98 identified the LIM homeobox 4 (LHX4) gene as highly enriched, along with known TFs in 99 this population such as THRB, ONECUT1, and OTX2. This transcript has significant fold 100 change (b = 3.3) and a low number of reads in the non-ThrbCRM1 active population, 101suggesting high specificity towards the cone/HC RPC population at this time (Fig 1 B). 102 A previous report has examined the presence of LIM-domain factors in early 103 chick photoreceptor development 15 . This study suggested that LHX3 was abundantly 104 present in the apical portion of the retina and localized to photoreceptors once the ONL 105 is clearly distinguished. As the RNA-Seq data indicated that LHX4 expression is 106 prominent in ThrbCRM1 reporter-positive cells at early stages while LHX3 transcript 107presence is marginal in all targeted cells (Supp. Fig. 1 A), we suspected that this 108 previous study could have detected LHX4 instead of LHX3 at earlier timepoints. To test 109 this, we electroporated a mouse LHX4 misex...

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“…To demonstrate the use of our dataset as a benchmarking reference, we compared the scRNA-seq profiles of distinct cell types generated using alternative methods, including fetal human cone photoreceptors, human induced pluripotent stem cell derived-cone photoreceptors (hiPSC-cone; (Welby et al, 2017), and a sample of adult human retina with 139 cells (Phillips et al, 2018). Correlation analysis demonstrated that the adult human retina sample showed highest similarity to rod photoreceptor (0.63, Supplementary figure 10), which is expected as rod photoreceptors represent the majority of the cells in the retina.…”

Section: Using the Human Neural Retina Transcriptome Atlas For Benchmmentioning

confidence: 99%

“…Phillips et al have profiled a total of 139 adult retina cells using the C1 Fluidigm platform (Phillips et al, 2018), but the limited number of profiled cells presents challenges in the annotation and accurate identification of individual retinal cell types. Moreover, a flow cytometry approach was used to isolate 65 human fetal cone photoreceptors followed by scRNA-seq profiling (Welby et al, 2017).…”

Section: Introductionmentioning

confidence: 99%

Generation of human neural retina transcriptome atlas by single cell RNA sequencing

Lukowski

Sharov

et al. 2018

Preprint

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The retina is a highly specialized neural tissue that senses light and initiates image processing. Although the functional organisation of specific cells within the retina has been well-studied, the molecular profile of many cell types remains unclear in humans. To comprehensively profile cell types in the human retina, we performed single cell RNA-sequencing on 20,009 cells obtained post-mortem from three donors and compiled a reference transcriptome atlas. Using unsupervised clustering analysis, we identified 18 transcriptionally distinct clusters representing all known retinal cells: rod photoreceptors, cone photoreceptors, Müller glia cells, bipolar cells, amacrine cells, retinal ganglion cells, horizontal cells, retinal astrocytes and microglia. Notably, our data captured molecular profiles for healthy and early degenerating rod photoreceptors, and revealed a novel role of MALAT1 in putative rod degeneration. We also demonstrated the use of this retina transcriptome atlas to benchmark pluripotent stem cell-derived cone photoreceptors and an adult Müller glia cell line. This work provides an important reference with unprecedented insights into the transcriptional landscape of human retinal cells, which is fundamental to our understanding of retinal biology and disease.

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Isolation and Comparative Transcriptome Analysis of Human Fetal and iPSC-Derived Cone Photoreceptor Cells

Cited by 96 publications

References 51 publications

A single‐cell transcriptome atlas of the adult human retina

A single‐cell transcriptome atlas of the adult human retina

Identification of genes with enriched expression in early developing mouse cone photoreceptors

Generation of human neural retina transcriptome atlas by single cell RNA sequencing

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