CRISPRi DSP-kd in iAT2
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ABSTRACT:
PROVIDER: PRJNA781443 | ENA |
REPOSITORIES: ENA
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Project description:We performed single cell transcriptomic profiling of induced human pluripotent stem cells (iPSCs)-derived type 2 alveolar epithelial cells (iAT2). iPSCs stably expressed CRISPRi (dCas9-KRAB) under the control of doxycyline. iAT2s were transduced with a lentivirus expressing gRNA targeting the transcriptional start site of DSP. Cells were treated with or without doxycyline to intiate CRISPRi-knockdown. Prior to capture, cells were labelled with hashing antibodies (HTO). Cells were captured for 10x Genomics Single cell capture (V3 protocol), and GEX and HTO libraries were sequenced. HTODemux function was used to demultiplex the samples. Knockdown of DSP caused cells to cluster separately from wild-type cells.
2022-05-26 | GSE189068 | GEO
Project description:Single-cell RNA-seq of CRISPRi ADGRG6-kd in iAT2
| PRJNA924804 | ENA
Project description:We performed single cell transcriptomic profiling of induced human pluripotent stem cells (iPSCs)-derived type 2 alveolar epithelial cells (iAT2). iPSCs stably expressed CRISPRi (dCas9-KRAB) under the control of doxycyline. iAT2s were transduced with a lentivirus expressing gRNA targeting the transcriptional start site of ADGRG6. Cells were treated with or without doxycyline to intiate CRISPRi-knockdown. Prior to capture, cells were labelled with hashing antibodies (HTO). Cells were captured for 10x Genomics Single cell capture (V3 protocol), and GEX and HTO libraries were sequenced. HTODemux function was used to demultiplex the samples. Knockdown of ADGRG6 caused cells to cluster separately from wild-type cells.
2023-09-20 | GSE223078 | GEO
Project description:We performed bulk transcriptomic profiling of induced human pluripotent stem cells (iPSCs)-derived type 2 alveolar epithelial cells (iAT2). iPSCs stably expressed CRISPRi (dCas9-KRAB) under the control of doxycyline. iAT2s were transduced with a lentivirus expressing gRNA targeting the transcriptional start site of ADGRG6. Cells were treated with or without doxycyline to intiate CRISPRi-knockdown. Cells were plated at an air-liquid interface, then subsequently exposed to air or 5% cigarette smoke using a VitroCell smoke robot. Cells were harvested for bulk RNA sequencing 8 hours post cigarette smoke exposure
2023-09-20 | GSE223077 | GEO
Project description:Bulk RNA-seq of CRISPRi ADGRG6-kd iAT2 following cigarette smoke exposure
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Project description:CBFB-DSP-ChIPseq and YAP-DSP-ChIPseq
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Project description:We developed a new method to extract open-chromatins that contain trans-acting factors (TAFs)-DNA complex crosslinked with dithiobis(succinimidyl propionate) (DSP). Using this method we identified CBFB as a cell density-sensitive TAF, which translocates between the cytoplasm and the nucleus in cell density-dependent manner. Here, we demonstrate that the extracted TAF/DNA complex is also amenable to ChIP-seq analysis.
2023-12-05 | GSE234731 | GEO
Project description:To further characterized the serrated pathway, we used the novel Digital Spatial Profiling (DSP) technology and its mRNA Cancer Transcriptome Atlas (CTA) panel, which includes over 1800 target gene, to investigate the underlying gene expression changes and pathways involved in sessile serrated lesion (SSL) and traditional serrated adenoma (TSA), two precancerous lesions to carcinoma in serrated pathway.
2023-12-31 | E-MTAB-12465 | biostudies-arrayexpress
Project description:Introduction: Glioblastoma (GBM) invasion studies have focused on coding genes, while few studies evaluate long non-coding RNAs (lncRNAs), transcripts without protein-coding potential, for role in GBM invasion. We leveraged CRISPR-interference (CRISPRi) to evaluate invasive function of GBM-associated lncRNAs in an unbiased functional screen, characterizing and exploring the mechanism of identified candidates. Methods: We implemented a CRISPRi lncRNA loss-of-function screen evaluating association of lncRNA knockdown (KD) with invasion capacity in Matrigel. Top screen candidates were validated using CRISPRi and oligonucleotide(ASO)-mediated knockdown in three tumor lines. Clinical relevance of candidates was assessed via The Cancer Genome Atlas(TCGA) and Genotype-Tissue Expression(GTEx) survival analysis. Mediators of lncRNA effect were identified via differential expression analysis following lncRNA KD and assessed for tumor invasion using knockdown and rescue experiments. Results: Forty-eight lncRNAs were significantly associated with 33-83% decrease in invasion (p<0.01) upon knockdown. The top candidate, LINC03045, identified from effect size and p-value, demonstrated 82.7% decrease in tumor cell invasion upon knockdown, while LINC03045 expression was significantly associated with patient survival and tumor grade(p<0.0001). RNAseq analysis of LINC03045 knockdown revealed that WASF3, previously implicated in tumor invasion studies, was highly correlated with lncRNA expression, while WASF3 KD was associated with significant decrease in invasion. Finally, WASF3 overexpression demonstrated rescue of invasive function lost with LINC03045 KD. Conclusion: CRISPRi screening identified LINC03045, a previously unannotated lncRNA, as critical to GBM invasion. Gene expression is significantly associated with tumor grade and survival. RNA-seq and mechanistic studies suggest that this novel lncRNA may regulate invasion via WASF3.
2024-06-03 | GSE268342 | GEO