Mus musculus
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA382181 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR5441490_1.fastq.gz | Fastqsanger.gz | |||
| SRR5441490_2.fastq.gz | Fastqsanger.gz | |||
| SRR5441491_1.fastq.gz | Fastqsanger.gz | |||
| SRR5441491_2.fastq.gz | Fastqsanger.gz | |||
| SRR5441492_1.fastq.gz | Fastqsanger.gz |
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Project description:RNA sequencing of developing cone photoreceptors
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Project description:RNA sequencing and ATAC sequencing of developing cone photoreceptors
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Project description:The loss of cone photoreceptor cells, which are critical for optimal daylight vision, have the great impact on vision during retinal degenerations. Retinal differentiation of human induced pluripotent stem cell (hiPSC) sources could provide a renewable source of cone photoreceptors towards developing a cone cell replacement therapy to treat blindness. Demonstration of comparable gene expression profiles between human foetal and stem cell-derived cones at equivalent stages is required to progress the cell transplantation approach into the patient, as it is hypothesised stem cell-derived cones are required to show high levels of developmental recapitulation of the in vivo generated cones. In this study, the AAV2/9.pR2.1.GFP reporter was used to specifically label L/M-opsin cone photoreceptors in human foetal retinal samples, obtained from the MRC-Wellcome Trust Human Developmental Biology Resource, at a range of developmental stages. The L/M-opsin cone population represent the majority cone cell types in the adult human retina and are the only photoreceptors present within the fovea. Using fluorescence activated cell sorting, L/M-opsin GFP+ cones and GFP- retinal populations, alongside total foetal retinal samples containing all retinal cell tytpes, were isolated and processed for bulk RNA sequencing and downstream comparative analysis. Using DESeq2 differential gene expression analyses, statistically significant genes enriched within the GFP+ human foetal LM-opsin cone populations were determined which led to the identification of a cone enriched gene signature of human L/M-opsin cone photoreceptors. The AAV2/9.pR2.1.GFP reporter was applied to hiPSC-derived retinal cultures to isolate and process cone-like cell populations for RNA sequencing using the same strategy developed within the human foetal retina. Applying the cone enriched gene signature to the transcriptome of hiPSC-derived GFP+ samples at equivalent developmental stages revealed some expression similarities in genes found to be enriched within the late foetal L/M-opsin cone photoreceptors. This analysis overall revealed an intermediate stage of cone differentiation was achieved within the hiPSC-derived samples and the comparison to human foetal L/M-opsin gene express profiles suggesting further differentiation of hiPSC-derived sample is required.
2018-04-20 | E-MTAB-6057 | biostudies-arrayexpress
Project description:Human cone photoreceptors represent a rare cell population of the human retina and are largely outnumbered by other retinal cell types. Despite their rarity, cone photoreceptors are critical for optimal daylight vision and their loss has the greatest impact on sight during the progression of retinal dystrophies. This is most applicable to the L/M-opsin cone populations which form the largest proportion of cones in the human retina and are the only cone types present within the fovea, which is responsible for fine detailed vision. Better understanding of the early genesis of these cells within the human developing retina, particularly in relation to the expression of genes that could be involved in regulating early events such as cell fate decision, is required. These molecular determinants could be exploited in vitro to generate more cone photoreceptors within human stem cell-derived retinal differentiation culture systems, suitable for use in cell transpantation therapies or disease modelling purposes. Additionally, the nature of this cell population in terms of their gene expression heterogeneity remains to be defined. State of the art technology has now advanced so that mRNA sequencing can be performed on individual cells, which means the transcriptome of rare cell populations such as cone photoreceptor cells can be interrogated without the need for ample material. Using the AAV2/9 pR2.1:GFP reporter, containing a synthetic promoter region previously characterised to drive expression in cone photoreceptors, we label and isolate human foetal L/M-opsin cone photoreceptors within the 15pcw human retinal retina. Isolated GFP+ cells were separated into individual cells using the Fluidigm C1 microfludics system, before RNA extraction and cDNA library preparation was performed. In total, 74 single cells were captured and processed for RNA sequencing; quality control of the data led to the inclusion of 65 individual cells for the downstream analysis. Principal component analysis revealed a very subtle heterogeneity across the cells, with the highest variability observed according to the first principal component (PC1). Differential gene expression analysis correlated to PC1 led to the identification of 503 significantly differentially expressed genes. Notably, some genes increase in expression across the cells when ordered according to PC1 which have previous associations with photoreceptor maturation, whereas other genes associated with developmental processes became downregulated across the cells. This led to the hypothesis that at this single timepoint in the human foetal retina, developing L/M-opsin cone photoreceptors exist at different stages of maturation. Overlapping genes were identified from comparing RNA seq data from 15pcw single cells and population samples at early and late timepoints of pR2.1:GFP+ cells, which showed the same gene expression trend between experiments. This suggests differences in cell maturation at a single timepoint of development could be representative of their true developmental trajectory. This overlapping gene dataset is also highly useful to define new gene that could be involved in cone photoreceptor development.
2018-04-20 | E-MTAB-6058 | biostudies-arrayexpress
Project description:Introgressed variants from other species can be an important source of genetic variation because they may arise rapidly, can include multiple mutations on a single haplotype, and have often been pretested by selection in the species of origin. Although introgressed alleles are generally deleterious, several studies have reported introgression as the source of adaptive alleles-including the rodenticide-resistant variant of Vkorc1 that introgressed from Mus spretus into European populations of Mus musculus domesticus. Here, we conducted bidirectional genome scans to characterize introgressed regions into one wild population of M. spretus from Spain and three wild populations of M. m. domesticus from France, Germany, and Iran. Despite the fact that these species show considerable intrinsic postzygotic reproductive isolation, introgression was observed in all individuals, including in the M. musculus reference genome (GRCm38). Mus spretus individuals had a greater proportion of introgression compared with M. m. domesticus, and within M. m. domesticus, the proportion of introgression decreased with geographic distance from the area of sympatry. Introgression was observed on all autosomes for both species, but not on the X-chromosome in M. m. domesticus, consistent with known X-linked hybrid sterility and inviability genes that have been mapped to the M. spretus X-chromosome. Tract lengths were generally short with a few outliers of up to 2.7 Mb. Interestingly, the longest introgressed tracts were in olfactory receptor regions, and introgressed tracts were significantly enriched for olfactory receptor genes in both species, suggesting that introgression may be a source of functional novelty even between species with high barriers to gene flow.
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Project description:We determined the accessible genome of postmitotic cone photoreceptors in the central region of the mouse retina on postnatal days P3, P6, and P10. At eachtimepoint we isolated GFP-labeled cells from three different Chrnb4-GFP mice (biological triplicates) using fluorescence-activated cell sorting. We then acquired reads of regions of accessible DNA of the sorted cones using ATAC sequencing. Our data set contained 9 samples.
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Project description:In retinal degenerative diseases, including retinitis pigmentosa (RP) and age-related macular degeneration (AMD), loss of photoreceptors at the macula leads to blindness. Among photoreceptors, rod cells are more vulnerable to degenerative stress than cone cells. Following disruption of neural retina leucine zipper (NRL) expression induced by knockout or the CRISPR/Cas9 system, rod cells have been shown to gain increased expression of several cone cell markers and demonstrate improved survival under degenerative pressure, consequently preventing secondary cone cell degeneration. Here, we report an mRNA silencing approach in which a cell-penetrating asymmetric small interfering RNA (cp-asiRNA) targeting NRL (cp-asiNRL) in postmitotic photoreceptors was used to suppress the expression of the NRL, which determines the fate of developing photoreceptors to rod cells. In mouse models of RP and neovascular AMD, intravitreal delivery of cp-asiNRL substantially increased the expression of cone-specific genes among the photoreceptor population and improved rod cell survival. Furthermore, in retinal organoids, cp-asiNRL also increased the production of photoreceptors expressing cone-specific markers. Our data suggest that cp-asiNRL-mediated NRL suppression in rod cells may be a promising strategy for treating retinal degenerative diseases.
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