RNA sequencing of E15.5 retinal tissue from control and Lhx2 CKO mice
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA723339 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR14289015.fastq.gz | Fastqsanger.gz | |||
| SRR14289016.fastq.gz | Fastqsanger.gz | |||
| SRR14289017.fastq.gz | Fastqsanger.gz | |||
| SRR14289018.fastq.gz | Fastqsanger.gz | |||
| SRR14289019.fastq.gz | Fastqsanger.gz |
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Project description:Lhx2 is a retinal progenitor cell transcription factor critical for eye development. We previously reported that conditional inactivation of Lhx2 at the start of mouse retinal neurogenesis disrupted retinal progenitor cell (RPC) proliferation, greatly reduced the RPC pool and altered neurogenic output as indicated by changes in the production of multiple fated precursor populations. To identify genes whose expression levels are dependent on Lhx2 at this stage of development, Lhx2 conditional inactivation was initiated at E11.5 in RPCs with the progenitor Cre driver Hes1CreERT2 and retinal tissue was collected at E15.5 for RNA sequencing. The gene expression profiles of Lhx2 CKO retinas were compared to control (Lhx2 conditional heterozygotes) were compared. Downregulated and upregulated gene expression was observed, with some likely due to direct and indirect regulation by Lhx2 within RPCs and others due to changes in differentiation and the altered neurogenic output.
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Project description:We profiled retinal progenitor cells by integrating next generation sequencing methods and interrogating changes in chromatin accessibility at embryonic and post-natal murine stages. We identified putative factors involved in the developmental progression of the retinal progenitors epigenome and we report Lhx2 role in the regulation of chromatin accessibility by coordinated action of developmentally regulated pioneer factors
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Project description:Eye formation is regulated by a complex network of eye field transcription factors (EFTFs), including LIM-homeodomain gene LHX2. We disrupted LHX2 function at different stages during this process using a conditional knock-out strategy in mice. We find that LHX2 function is required in an ongoing fashion to maintain optic identity across multiple stages, from the formation of the optic vesicle to the differentiation of the neuroretina. At each stage, loss of Lhx2 led to upregulation of a set of molecular markers that are normally expressed in the thalamic eminence and in the anterodorsal hypothalamus in a portion of the optic vesicle or retina. Furthermore, the longer LHX2 function was maintained, the further optic morphogenesis progressed. Early loss of function caused profound mispatterning of the entire telencephalic-optic-hypothalamic field, such that the optic vesicle became mispositioned and appeared to arise from the diencephalic-telencephalic boundary. At subsequent stages, loss of Lhx2 did not affect optic vesicle position but caused arrest of optic cup formation. If Lhx2 was selectively disrupted in the neuroretina from E11.5, the neuroretina showed gross dysmorphology along with aberrant expression of markers specific to the thalamic eminence and anterodorsal hypothalamus. Our findings indicate a continual requirement for LHX2 throughout the early stages of optic development, not only to maintain optic identity by suppressing alternative fates but also to mediate multiple steps of optic morphogenesis. These findings provide new insight into the anophthalmic phenotype of the Lhx2 mutant and reveal novel roles for this transcription factor in eye development.
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Project description:Tissue-specific transcription factors control the transcriptome through an association with noncoding regulatory regions (cistromes). Identifying the combination of transcription factors that dictate specific cell fate, their specific cistromes and examining their involvement in complex human traits remain a major challenge. Here we focus on the retinal pigmented epithelium (RPE), an essential lineage for retinal development and function and the primary tissue affected in age-related macular degeneration (AMD), a leading cause of blindness. By combining mechanistic findings in stem-cell-derived human RPE, in- vivo functional studies in mice and global transcriptomic and proteomic analyses, we revealed that the key developmental transcription factors LHX2 and OTX2 function together in transcriptional module containing LDB1 and SWI/SNF (BAF) to regulate the RPE transcriptome. Importantly, the intersection between the identified LHX2-OTX2 cistrome with published expression quantitative trait loci, ATAC-seq data from human RPE, and AMD GWAS data, followed by functional validation using a reporter assay, revealed a causal genetic variant that affects AMD risk by altering TRPM1 expression in the RPE through modulation of LHX2 transcriptional activity on its promoter. Taken together, the reported cistrome of LHX2 and OTX2, the identified downstream genes and interacting co-factors reveal the RPE transcription module and uncover a causal regulatory risk SNP in the multifactorial common blinding disease AMD.
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Project description:RNA-seq analysis was applied in triplicates to RPE cells extracted at two developmental time-points (E15.5 and P5) from Sox9-cKO and wild-type control mice
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Project description:Purpose: Müller glia are the only glial cell type produced by the neuroepithelial progenitor cells which generate the vertebrate retina. Müller glia are required to maintain retinal homeostasis and support the survival of retinal neurons. Furthermore, they function as an adult stem cell, mediating retinal regeneration among select vertebrate classes. However, the mechanisms which regulate Müller development are poorly understood as considerable overlap exists in gene expression between retinal progenitor cells and differentiated Müller glia. We investigate the functional role of the LIM homeodomain transcription factor Lhx2 in the specification and development of Müller glia in the mouse. Methods: RNA-Seq was performed in collaboration with the Johns Hopkins School of Medicine Deep Sequencing and Microarray Core Facility. Libraries were prepared using Illumina TruSeq RNA Sample kit (Illumina, San Diego, CA) following manufacturerâs recommended procedure. The PCR amplified library was purified using RNAClean XP magnetic beads (Agencourt, Beverley, MA) and run out on a High Sensitivity DNA Chip (Agilent, Santa Clara, CA) for quality check. We used STAR to align RNA-Seq reads onto Ensembl mouse genome GRCm38, release 72. To generate the stand attribute for alignments containing splice junctions, we used the outSAMstrandField intronMotif program. The spliced alignments without strand definition were removed. Number of reads mapped to exons was counted by htseq-count. Genes expressed at very low levels were omitted from further analysis. Gene expression differences between wildtype and mutant samples, significance (p-value) and false discovery rate (FDR) were computed using the generalized linear models based EdgeR. Results: We observed a substantial reduction in expression of Notch pathway genes including Notch1, the Notch ligands Dll1 and Dll3, as well as gliogenic Notch effector genes such as Hes1, Hes5, Id1 and Sox8 and the Müller-gliogenic factor Rax. We likewise observe a substantial reduction in expression of progenitor-specific genes such as Vsx2 and Fgf15. Furthermore, we observed a decrease in the expression of early-onset glial markers such as Crym , Spon1, and Car2. Retinal mRNA profiles of post-natal day 0.5 (P0.5) Lhx2 wild type (N=3) and Lhx2lox/lox; Pdgfrα-Cre ÎcKO (N=3) mice were generated using Illumina TruSeq and analyzed with Agilent high sensitivity DNA analsis kit.
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Project description:We report RNAseq analysis of the transcriptome of retinas from mature rod-specific Dicer1 cKO mice and control littermates lacking Cre expression in order to better understand changes in gene regulation that could lead to retinal degeneration in cKO mice. Examine retinal transcriptome of 3 biological replicates for each genotype from 4-week-old animals with tissue collected between 8:00 - 10:00AM
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2014-08-14 | E-GEOD-55376 | biostudies-arrayexpress
Project description:Expression of the lim-homeodomain transcription factor is required for sustained maintenance of heamatopoietic stem cell like cells in undifferentiated form durng in vitro culturing. Cell lines were created from the mouse embryonic stem (ES) cell line, Ainv15. The gene Lhx2 was introducted into these cells under the control of a tetracycline-responsive element. In the presence of tetracycline (or its analogue doxycycline), these cells express Lhx2. In DoxHPC7 GFP is co-expressed together with the Lhx2 gene. We used these cells to carry out a time-course study where the effects of Lhx2 withdrawal were studied.
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