High-throughput RNA-sequencing-based transcriptomic profiles of embryonic lens development for cataract gene discovery
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE119596 | GEO | 2019/01/08
REPOSITORIES: GEO
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Project description:Identification of genes involved in ocular birth defects remains a challenge. To facilitate the identification of genes associated with cataract, we developed iSyTE (integrated Systems Tool for Eye gene discovery; http://bioinformatics.udel.edu/Research/iSyTE). iSyTE contains microarray gene expression profiles of the mouse embryonic lens as it transitions from the stage of placode invagination to that of vesicle formation. We identified differentially regulated genes by comparing lens microarray profiles to those representing whole embryonic body (WB) without ocular tissue. These were then utilized to generate a ranked list of lens-genes enrichment, which can be viewed as iSyTE tracks in the UCSC Genome browser to aid identification of genes with lens function. We microdissected embryonic lens from mice at E10.5, E11.5, and E12.5 (triplicate at each time point). To have a proper control, we also generated gene expression profiles of whole embryonic body (WB) at these time points. For comparative analysis, we also generated gene expression profiles of E13.5 tooth germs tissues, and matched WB. These profiles are used to identify lens and tooth specific gene expression enrichment at these embryonic time points. This dataset is then used to prioritize analysis of candidate cataract associated genes.
2012-02-16 | E-GEOD-32334 | biostudies-arrayexpress
Project description:Identification of genes involved in ocular birth defects remains a challenge. To facilitate the identification of genes associated with cataract, we developed iSyTE (integrated Systems Tool for Eye gene discovery; http://bioinformatics.udel.edu/Research/iSyTE). iSyTE contains microarray gene expression profiles of the mouse embryonic lens as it transitions from the stage of placode invagination to that of vesicle formation. We identified differentially regulated genes by comparing lens microarray profiles to those representing whole embryonic body (WB) without ocular tissue. These were then utilized to generate a ranked list of lens-genes enrichment, which can be viewed as iSyTE tracks in the UCSC Genome browser to aid identification of genes with lens function.
2012-02-16 | GSE32334 | GEO
Project description:Although majority of the genes linked to pediatric cataract exhibit lens fiber cell-enriched expression, our understanding of gene regulation in these cells is limited to function of just eight transcription factors and largely in the context of crystallins. Here, we identify small Maf transcription factors MafG and MafK as regulators of several non-crystallin human cataract genes in fiber cells and establish their significance to cataract. We applied a bioinformatics tool for cataract gene discovery iSyTE to identify MafG and its co-regulators in the lens, and generated various null-allelic combinations of MafG:MafK mouse mutants for phenotypic and molecular analysis. By age 4-months, MafG-/-:MafK+/- mutants exhibit lens defects that progressively develop into cataract. High-resolution phenotypic characterization of MafG-/-:MafK+/- lens reveals severe defects in fiber cells, while microarrays-based expression profiling identifies 97 differentially regulated genes (DRGs). Integrative analysis of MafG-/-:MafK+/- lens-DRGs with 1) binding-motifs and genomic targets of small Mafs and their regulatory partners, 2) iSyTE lens-expression data, and 3) interactions between DRGs in the String database, unravels a detailed small Maf regulatory network in the lens, several nodes of which are linked to human cataract. This analysis prioritizes 36 highly promising candidates from the original 97 DRGs. Significantly, 8/36 (22%) DRGs are associated with cataracts in human (GSTO1, MGST1, SC4MOL, UCHL1) or mouse (Aldh3a1, Crygf, Hspb1, Pcbd1), suggesting a multifactorial etiology that includes elevation of oxidative stress. These data identify MafG and MafK as new cataract-associated candidates and define their function in regulating largely non-crystallin genes linked to mouse and human cataract. Microarray comparision of lenses from mixed background (129Sv/J, C57BL/6J, and ICR) control (MafG+/-:MafK+/-; no-cataract) and compound (MafG-/-:MafK+/-; cataract) mouse mutants
2015-05-01 | E-GEOD-65500 | biostudies-arrayexpress
Project description:To expedite gene discovery in eye development and its associated defects, we previously developed a bioinformatics resource-tool iSyTE (integrated Systems Tool for Eye gene discovery). However, iSyTE is presently limited to lens tissue and is predominantly based on transcriptomics datasets. Therefore, to extend the iSyTE approach to retina development on the proteome level, we performed high-throughput tandem mass spectrometry (MS/MS) on mouse embryonic day (E)14.5 retina and identified an average of 3,300 proteins per sample (n=5). High-throughput expression profiling-based gene discovery approaches–involving either transcriptomics or proteomics–pose a key challenge of prioritizing select candidates from thousands of RNA/proteins expressed in a particular cell/tissue type. To address this, we used MS/MS proteome data from mouse whole embryonic body (WB) as a reference dataset and performed comparative analysis–termed “in silico WB-subtraction”–with the retina proteome dataset. In silico WB-subtraction identified 90 high-priority proteins with retina-enriched expression at stringency criteria of 2.5 average spectral counts, 2.0 fold-enrichment, False Discovery Rate <0.01. These top candidates represent a pool of retina-enriched proteins, several of which are associated with retinal biology and/or defects (e.g., Aldh1a1, Ank2, Ank3, Dcn, Dync2h1, Egfr, Ephb2, Fbln5, Fbn2, Hras, Igf2bp1, Msi1, Rbp1, Rlbp1, Tenm3, Yap1, etc.), indicating the effectiveness of this approach. Importantly, in silico WB-subtraction also identified several new high-priority candidates with potential regulatory function in retina development. Finally, proteins exhibiting expression or enriched-expression in the retina are made accessible in a user-friendly manner at iSyTE (https://research.bioinformatics.udel.edu/iSyTE/), to allow effective visualization of this information by the research community to facilitate eye gene discovery.
2023-10-09 | PXD039490 | Pride
Project description:Although majority of the genes linked to pediatric cataract exhibit lens fiber cell-enriched expression, our understanding of gene regulation in these cells is limited to function of just eight transcription factors and largely in the context of crystallins. Here, we identify small Maf transcription factors MafG and MafK as regulators of several non-crystallin human cataract genes in fiber cells and establish their significance to cataract. We applied a bioinformatics tool for cataract gene discovery iSyTE to identify MafG and its co-regulators in the lens, and generated various null-allelic combinations of MafG:MafK mouse mutants for phenotypic and molecular analysis. By age 4-months, MafG-/-:MafK+/- mutants exhibit lens defects that progressively develop into cataract. High-resolution phenotypic characterization of MafG-/-:MafK+/- lens reveals severe defects in fiber cells, while microarrays-based expression profiling identifies 97 differentially regulated genes (DRGs). Integrative analysis of MafG-/-:MafK+/- lens-DRGs with 1) binding-motifs and genomic targets of small Mafs and their regulatory partners, 2) iSyTE lens-expression data, and 3) interactions between DRGs in the String database, unravels a detailed small Maf regulatory network in the lens, several nodes of which are linked to human cataract. This analysis prioritizes 36 highly promising candidates from the original 97 DRGs. Significantly, 8/36 (22%) DRGs are associated with cataracts in human (GSTO1, MGST1, SC4MOL, UCHL1) or mouse (Aldh3a1, Crygf, Hspb1, Pcbd1), suggesting a multifactorial etiology that includes elevation of oxidative stress. These data identify MafG and MafK as new cataract-associated candidates and define their function in regulating largely non-crystallin genes linked to mouse and human cataract.
2015-05-01 | GSE65500 | GEO
Project description:Purpose: While the bioinformatics gene-discovery tool iSyTE (integrated Systems Tool for Eye gene discovery) has proved effective in identifying new cataract-linked genes, it primarily depended on lens-enriched expression of just three wild-type mouse embryonic stages. To increase iSyTE’s efficacy, a broad range of embryonic (E) and postnatal (P) lens microarray datasets from wild-type and specific gene-perturbation mutant mice were analyzed and a new web-interface was developed for their effective access and downstream-analysis. Methods: Five new mouse lens microarrays were generated and analyzed with all publicly available Affymetrix and Illumina lens datasets representing the stages E9.5, E10.25, E10.5, E11.5, E12.5, E15.5, E16.5, E17.5, E19.5, P0, P2, P4, P8, P12, P20, P28, P30, P42, P52, P56, P60 and mouse mutants for the genes: Brg1, CBP:p300, E2f1:E2f2:E2f3, Foxe3, Hsf4, Klf4, Mafg:Mafk, Notch2, Pax6, Sparc, Tdrd7. Results: The integrated lens microarray analysis accurately reflects established gene expression patterns and defines a new lens-signature gene-set for embryonic and postnatal stages that includes several novel candidates namely, Grifin, Ogn, Fabp5, Mboat1, Tmem40, Dhx32, Aldoc, Pgam2, Hmgn3, Mocs2, Gprc5b, Gstm1, Npl, and Zbtb8b. Combined analyses of 32 developmental/genetic data-conditions identifies several new transcription and signaling factors that potentially function in lens development and/or homeostasis. Conclusions: The updated iSyTE web-interface: (1) comprehensively informs on normal lens developmental and cataract-associated transcriptome dynamics and identifies new high-priority candidates for these processes, and (2) offers user-friendly visualization.
2017-09-21 | GSE100136 | GEO
Project description:The Shumiya cataract rat (SCR) is a model for hereditary cataract. Two-third of these rats develop lens opacity within 10-11-weeks. Onset of cataract is attributed to the synergetic effect of lanosterol synthase (Lss) and farnesyl-diphosphate farnesyltransferase 1 (Fdft1) mutant alleles that lead to cholesterol deficiency in the lenses, which in turn adversely affects lens biology including the growth and differentiation of lens epithelial cells (LECs). Nevertheless, the molecular events and changes in gene expression associated with the onset of lens opacity in SCR is poorly understood. Our study aimed to identify the gene expression patterns during cataract formation in SCRs, which may be responsible for cataractogenesis in SCR.
2020-07-31 | GSE152616 | GEO
Project description:Mutations of the RNA-granule component TDRD7 (OMIM: 611258) cause pediatric cataract in humans. Here, we applied an integrated approach to elucidate the molecular pathology of cataract in Tdrd7 targeted-knockout (Tdrd7-/-) mice. Tdrd7-/- animals precipitously develop lens fiber cell abnormalities early in life, suggesting a global-level breakdown/mis-regulation of key cellular processes. High-throughput RNA-sequencing followed by iSyTE-integrated bioinformatics-based analysis identified the molecular chaperone and cytoskeletal-modulator, HSPB1 (HSP27), among the high-priority down-regulated candidates in Tdrd7-/- lens. Moreover, a protein 2-D fluorescence difference gel electrophoresis-coupled mass spectrometry screen also identified HSPB1 to be reduced in Tdrd7-/- lens, offering independent support for focusing efforts on this factor to explain Tdrd7-/- cataract. Reduction of HSPB1 preceded lens morphological abnormalities, suggesting that cytoskeletal defects underlie the Tdrd7-/- cataract phenotype. In agreement, scanning electron microscopy revealed abnormal fiber cell membrane protrusions in Tdrd7-/- lenses. Significantly, abnormal F-actin staining was detected specifically in Tdrd7-/- fiber cells that exhibit nuclear degradation, thereby revealing that there are distinct mechanisms based on pre- or post-nuclear degradation differentiation stage for F-actin cytoskeletal maintenance in fiber cells. Further, RNA-immunoprecipitation identified Hspb1 mRNA in wild-type lens lysate TDRD7-pulldowns, and single-molecule RNA-imaging showed co-localization of TDRD7 protein with cytoplasmic Hspb1 mRNA in a specific pre-nuclear degradation area of differentiating fiber cells, indicating that TDRD7-ribonucleoprotein complexes are necessary for controlling optimal levels of key factors in lens development. Together, these data uncover a novel role for TDRD7 in regulating elevation of stress-responsive chaperones for cytoskeletal maintenance in post-nuclear degradation lens fiber cells, perturbation of which causes early-onset cataracts.
2020-09-01 | GSE134384 | GEO
Project description:Early changes in the transcriptome associated with lens wounding in an ex vivo post-cataract surgery chicken model. Here we report the changes in the transcriptome that occur 1hr vs. time 0 post-cataract surgery wounding. Our data provide a molecular framework for understanding the early gene changes associated with the injury response of the lens.
2023-11-04 | GSE239368 | GEO
Project description:Purpose: TGF-β/BMP signaling pathway has a significant role in fibrotic cataract. Smurf1, a ubiquitin protein ligase, regulates the TGF-β/BMP signaling pathway through the ubiquitin-proteasome system (UPS). This study aims to investigate the role of Smurf1 in the progression of fibrotic cataract and its underlying mechanism.Method: We employed a mouse injury-induced anterior subcapsular cataract (ASC) model and administered Smurf1 inhibitor A01 through anterior chamber injection for in vivo investigation. RNA sequencing was performed to examine global gene expression changes. The volume of the subcapsular opacity was determined using whole-mount immunofluorescence of lens anterior capsules. Lentivirus was utilized to create cell lines with Smurf1 knockdown or overexpression in SRA01/04. Protein levels were assessed by Simple Western. Lens epithelial cell (LEC) proliferation was evaluated by CCK8 and EdU assays. LEC migration was measured by Transwell and wound healing assays.Results: The mRNA levels of genes associated with cell proliferation, migration, epithelial-mesenchymal transition (EMT), TGF-β/BMP pathway and UPS, including Smurf1, were upregulated in ASC model. The mRNA expression of Smurf1 was also upregulated in anterior lens capsules of age-related cataract patients. Anterior chamber injection of A01 inhibited ASC formation and EMT. In vitro knockdown of Smurf1 resulted in reduced proliferation, TGF-β2-induced migration and EMT of LECs. Smurf1 inhibition upregulated Smad1, Smad5 and pSmad1/5. Conversely, Smurf1 overexpression displayed the opposite phenotypes.Conclusion: Smurf1 regulated the progression of fibrotic cataract by influencing the proliferation, migration and EMT of LECs through regulation of Smad signaling pathway, offering a novel target for the treatment of fibrotic cataract.
2024-02-14 | GSE245411 | GEO