The GPI Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HAP1 human near-haploid cell line, engineered to disrupt the GPI gene. This product provides a loss-of-function model for studying glucose-6-phosphate isomerase (GPI), a dual-function protein that serves as a glycolytic enzyme and as the secreted autocrine motility factor (AMF). The polyclonal knockout pool enables investigation of GPI-dependent processes without clonal selection, maintaining genetic heterogeneity that can be advantageous for functional genomics and drug discovery screens.
HAP1 cells are a fibroblast-like cell line with a near-haploid karyotype, originally derived from the KBM-7 chronic myeloid leukemia line. Their haploid nature facilitates CRISPR/Cas9-mediated gene disruption, as a single targeting event can produce a complete knockout, making them a robust model for loss-of-function studies. HAP1 cells are widely employed in functional genomics, cancer research, and drug target validation due to their genetic tractability and stable phenotype.
GPI catalyzes the reversible isomerization of glucose-6-phosphate to fructose-6-phosphate, a rate-limiting step in glycolysis that also feeds into the pentose phosphate pathway and gluconeogenesis. Beyond metabolism, secreted GPI (AMF) binds to the gp78/AMFR receptor, activating downstream signaling cascades including PI3K/AKT, ERK1/2, and Rho family GTPases. This signaling promotes expression of matrix metalloproteinases (MMPs) and enhances cell migration, invasion, and survival. GPI expression is regulated by HIF-1?? under hypoxia, and by growth factors (EGF, HGF) and oncogenic Ras, linking metabolic status to motility signaling.
In the HAP1 near-haploid background, GPI knockout disrupts both glycolytic flux and AMF signaling, providing a unique platform to dissect the interplay between metabolism and cell motility. This model enables researchers to study how loss of GPI impacts energy production, nucleotide biosynthesis via the pentose phosphate pathway, and the autocrine regulation of migration. It is particularly relevant for investigating mechanisms of cancer metastasis, metabolic adaptations in leukemia cells, and the role of AMF in immune cell recruitment in autoimmune diseases.
Typical experimental applications include analyzing glycolytic flux using Seahorse assays, assessing cell migration and invasion in Boyden chamber or wound healing assays, and measuring MMP activity by zymography. The polyclonal knockout can be used to validate GPI as a therapeutic target for anti-metastatic strategies, to study GPI deficiency-related hemolytic anemia, and to screen for small molecules that modulate AMF-gp78 interactions. Downstream signaling can be monitored via phospho-AKT/ERK western blotting, and AMF receptor binding assessed by immunoprecipitation. For more information on this product, please contact Ascent Research.
