Generation of Endogenous BMP Transcriptional Reporter Cells Through CRISPR/Cas9 Genome Editing

Abstract

Transcriptional reporter systems allow researchers to investigate the function and regulation of transcription factors. Conventional systems employ artificial cDNA overexpression vectors containing either a promoter fragment or specific nucleotide sequence repeats upstream of firefly luciferase or fluorescent reporters, such as green fluorescence protein (GFP) cDNA (1). These systems suffer mainly from the lack of chromatin context. Here, we describe the rapid generation of endogenous transcriptional reporter cells for the Bone Morphogenetic Protein (BMP) pathway using CRISPR/Cas9 genome editing. In principle, our methodology can be applied to any cell line. The endogenous reporters will provide a robust system for the investigation of BMP transcriptional activity in the context of native chromatin landscape and facilitate chemical and genetic screens.

Insights into Bone Morphogenetic Protein—(BMP-) Signaling in Ocular Lens Biology and Pathology

“…Freitas et al (2021) used CRISPR-Cas9 to overexpress BMP-9 in mesenchymal stem cells (MSCs) and when these genetically edited cells were injected into rat calvarial bone defects, the BMP-9-overexpressing MSCs were able to repair these defects, with increased bone formation and bone mineral density [158]. Hutchinson et al (2019) described an innovative methodology using CRISPR/Cas9 to generate endogenous transcriptional reporter cells for the BMP pathway, and this technique could be applied to ocular lens cells to enable future investigations of BMP transcriptional activity in lens development and pathology [159].…”