The IMPA2 Knockout HT29 Polyclonal Cells represent a CRISPR/Cas9-edited polyclonal knockout cell population derived from the human colorectal adenocarcinoma line HT29, engineered to disrupt the gene encoding inositol monophosphatase 2 (IMPA2). This pool, generated without clonal isolation, provides a heterogeneous loss-of-function model suited for bulk biochemical and functional analyses of IMPA2-dependent signaling.
HT29 cells are an adherent, epithelial-like cell line exhibiting microvilli characteristic of intestinal enterocytes, originally established from a primary colorectal tumor of a 44-year-old female. Extensively used to study intestinal absorption, drug transport, and colorectal cancer biology, this line offers a physiologically relevant platform for dissecting tumor-associated pathways and therapeutic responses.
IMPA2 catalyzes the dephosphorylation of inositol monophosphates to free myo-inositol, a critical step in recycling inositol for the synthesis of phosphatidylinositol and phosphoinositide second messengers. Its activity is inhibited by lithium and modulated by upstream regulators including TFAP2, SP1, CREB, and neuronal depolarization. Downstream targets encompass PIP2, IP3, DAG, PKC, and the prosurvival kinase AKT. By maintaining phosphoinositide pools, IMPA2 feeds into the phospholipase C?Ccalcium and PI3K-Akt pathways; disruption depletes IP3 and DAG, attenuating calcium mobilization and PKC activation, ultimately reducing AKT signaling. Key interacting partners include IMPA1, INPP1, IMPAD1, and lithium ion, with additional pathway components such as PI3K, PTEN, PLC, IP3 receptor, DAG kinase, ISYNA1, and MIOX.
In the colorectal cancer context of HT29 cells, IMPA2 knockout phenocopies lithium-induced inositol depletion, leading to impaired phosphoinositide recycling and blunted IP3/DAG?Ccalcium?CPKC cascades. The consequent reduction in AKT prosurvival signaling can suppress cell proliferation and enhance apoptotic sensitivity, offering a model to investigate the contribution of inositol metabolism to tumor growth, migration, and drug resistance, and to uncover targetable vulnerabilities.
These cells are well suited for applications spanning mechanistic studies of lithium??s impact on inositol signaling, high-throughput screening for modulators of phosphoinositide turnover, and functional genomics to identify compensatory networks. Typical assays include western blotting and RT-qPCR for expression analysis, HPLC- or mass spectrometry?Cbased inositol phosphate quantification, cell proliferation (MTT/BrdU) and apoptosis (Annexin V/7-AAD) assays, transwell migration/invasion, intracellular calcium flux (Fluo-4), phospho-AKT Ser473 ELISA, and colony formation. For further information, please contact Ascent Research.
