The ECE2 Knockout HAP1 Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HAP1 near-haploid human cell line, featuring targeted disruption of the endothelin-converting enzyme 2 (ECE2) gene. Unlike single-cell-derived clonal lines, this polyclonal format introduces a heterogeneous array of editing events across the cell pool, collectively abrogating ECE2 protein expression and enzymatic activity, making it a versatile tool for loss-of-function studies without the genetic bottleneck of clonal selection.
The HAP1 host cell line originates from the KBM-7 chronic myeloid leukemia (CML) line and retains a predominantly haploid karyotype (disomic only for chromosome 8), enabling straightforward gene disruption via targeting of a single allele. These adherent, fibroblast-like cells express the BCR-ABL fusion oncoprotein and serve as a robust model for genetic screens, cancer biology, and haploid genetics, combining the benefits of mammalian cell physiology with the genetic tractability of a haploid system.
ECE2 is a type II integral membrane metallopeptidase that cleaves big endothelin-1 to the vasoactive peptide endothelin-1. Endothelin-1 acts via ETA and ETB GPCRs, coupling to G??q/11 to stimulate PLC??, IP3-mediated calcium release, and DAG/PKC activation, while also engaging ERK1/2 and AKT pathways. ECE2 expression is driven by transcription factors GATA2 and SP1, and is enhanced by cytokines (IL-1??, TNF-??) and growth factors (TGF-??, EGF). It operates alongside ECE1 isoforms and is modulated by neprilysin, establishing a network for precise control of endothelin signaling.
In these knockout cells, disrupted ECE2 impedes endothelin-1 production, blunting downstream ERK1/2, AKT, and calcium signaling. The HAP1 CML background with BCR-ABL allows investigation of crosstalk between endothelin and oncogenic pathways, relevant to cancer cell proliferation and survival.
Applications include cardiovascular disease research, cancer biology, drug screening for ECE2 inhibitors, and functional genomics. Key assays: endothelin-1 ELISA, western blot (ECE2, phospho-ERK/AKT), RT-qPCR, cell proliferation, migration, and calcium flux. The polyclonal format is also suitable for phenotypic screens. For inquiries, contact Ascent Research.
