HHIPL2 Knockout HEK293T Polyclonal Cells are a CRISPR/Cas9-mediated gene disruption product consisting of a polyclonal population of HEK293T cells with targeted inactivation of the HHIPL2 gene. This loss-of-function model enables comprehensive examination of HHIPL2??s role in negative regulation of the hedgehog signaling pathway, without the need for single-cell cloning, thereby preserving population-level genetic heterogeneity relevant for pathway-modulation studies.
The HEK293T cell line originates from human embryonic kidney epithelium and has been immortalized with adenovirus type 5, additionally stably expressing the SV40 large T antigen. This enables high-copy episomal replication of plasmids containing an SV40 origin of replication, contributing to its remarkably high transfection efficiency. It is a widely adopted mammalian expression system for recombinant protein production, lentiviral packaging, and CRISPR-based engineering, offering an ideal background for generating knockout models of regulatory genes.
HHIPL2 encodes a secreted hedgehog-interacting protein that functions as an extracellular negative regulator of hedgehog signaling. The protein directly binds sonic hedgehog (SHH), Indian hedgehog (IHH), and desert hedgehog (DHH) ligands, preventing their interaction with the PTCH1 receptor. This sequestration blocks downstream activation of SMO and the GLI family of transcription factors (GLI1, GLI2, GLI3), ultimately dampening expression of target genes such as PTCH1, GLI1, and CCND1 that drive cell proliferation and differentiation. HHIPL2??s activity is modulated by interactions with heparan sulfate proteoglycans, including GPC3 and GPC6, which influence ligand distribution. Within the primary cilium-dependent signaling hub, HHIPL2 acts in concert with HHIP, KIF7, and SUFU to shape the gradient and dynamics of hedgehog responses.
HEK293T cells do not typically exhibit robust hedgehog pathway activity or ciliogenesis in standard culture, but their high transfectability allows facile introduction of pathway components or stimulation with recombinant ligands. Consequently, this HHIPL2 knockout polyclonal pool provides a clean, simplified system for dissecting the gene??s role in ligand sequestration and GLI-mediated transcription. By comparing wild-type and knockout cells upon pathway activation, researchers can quantify HHIPL2-dependent changes in signaling strength, reporter activity, and downstream gene expression, thereby clarifying its specific contribution to negative regulation without confounding endogenous ciliary signaling.
This product supports diverse research workflows, including functional analysis of hedgehog signaling in cancers such as basal cell carcinoma and medulloblastoma, developmental biology studies of morphogen gradients, and validation of small-molecule SMO or GLI inhibitors. Compatible assays comprise GLI luciferase reporter assays, RT-qPCR and western blotting for pathway components, co-immunoprecipitation of HHIPL2 with ligands, and cell proliferation assays (MTT/BrdU). When ciliogenesis is induced, immunofluorescence for primary cilia markers can be employed. For additional product specifications or to place an order, please contact Ascent Research.
