CRISPR/Cas9 engineering of a KIM-1 reporter human proximal tubule cell line.
Ruth Ann VeachMatthew H WilsonPublished in: PloS one (2018)
We used the CRISPR/Cas9 system to knock-in reporter transgenes at the kidney injury molecule-1 (KIM-1) locus and isolated human proximal tubule cell (HK-2) clones. PCR verified targeted knock-in of the luciferase and eGFP reporter at the KIM-1 locus. HK-2-KIM-1 reporter cells responded to various stimuli including hypoxia, cisplatin, and high glucose, indicative of upregulation of KIM-1 expression. We attempted using CRISPR/Cas9 to also engineer the KIM-1 reporter in telomerase-immortalized human RPTEC cells. However, these cells demonstrated an inability to undergo homologous recombination at the target locus. KIM-1-reporter human proximal tubular cells could be valuable tools in drug discovery for molecules inhibiting kidney injury. Additionally, our gene targeting strategy could be used in other cell lines to evaluate the biology of KIM-1 in vitro or in vivo.
Keyphrases
- crispr cas
- endothelial cells
- genome editing
- high glucose
- induced apoptosis
- cell cycle arrest
- signaling pathway
- induced pluripotent stem cells
- drug discovery
- pluripotent stem cells
- endoplasmic reticulum stress
- oxidative stress
- cell death
- cancer therapy
- transcription factor
- gene expression
- mesenchymal stem cells
- single cell
- dna methylation
- cell therapy