The AKT1S1 Knockout HAP1 Polyclonal Cells comprise a CRISPR/Cas9-edited polyclonal knockout cell population in which the AKT1S1 gene has been disrupted, generating a loss-of-function model for the encoded PRAS40 protein. This product provides a genetically heterogeneous pool of cells with targeted gene disruption, suitable for functional genomics studies without the clonal selection artifacts often associated with single-cell-derived lines.
HAP1 is a near-haploid human cell line originally derived from the KBM-7 chronic myeloid leukemia (CML) line. It exhibits a fibroblast-like morphology and grows in suspension, making it amenable to high-throughput screening and large-scale culture. The near-haploid karyotype simplifies genetic manipulation and enables unambiguous genotype?Cphenotype correlations, establishing HAP1 as a powerful model system for eukaryotic functional genomics and pathway dissection.
AKT1S1 encodes PRAS40 (proline-rich AKT substrate of 40 kDa), a key negative regulator of the mTORC1 (mechanistic target of rapamycin complex 1) signaling hub. In its unphosphorylated state, PRAS40 binds directly to RAPTOR, a component of mTORC1, and suppresses kinase activity. Upstream growth factor signals, such as insulin or IGF-1, activate the PI3K?CPDK1?CAKT cascade, leading to AKT-mediated phosphorylation of PRAS40 on threonine 246. This phosphorylation promotes 14-3-3 protein binding and dissociation of PRAS40 from mTORC1, thereby relieving inhibition and enabling mTORC1 to phosphorylate downstream targets including S6K1, 4E-BP1, and ULK1. Consequently, mTORC1 activation drives protein synthesis, ribosome biogenesis, cell cycle progression, and suppression of autophagy. PRAS40 thus integrates nutrient and growth factor cues to control cell growth and survival.
In the HAP1 CML background, disruption of AKT1S1 removes a critical brake on mTORC1 signaling, providing a unique platform to study sustained mTORC1 hyperactivity and its implications in leukemia biology and oncogenic transformation. Given that AKT1S1 is frequently dysregulated in solid tumors (e.g., breast, prostate) and hematological malignancies, this knockout model enables analysis of mTORC1-dependent proliferation and survival pathways without the confounding crosstalk typically present in diploid cancer lines. The haploid genetics further facilitate CRISPR-based modifier screens to identify synthetic lethal interactions or resistance mechanisms relevant to mTOR-targeted therapies.
Researchers can employ these polyclonal knockout cells in a wide array of applications, including signal transduction studies using phospho-specific antibodies for AKT (pS473) and S6K1 (pT389) by western blotting, mTORC1 activity assays, and co-immunoprecipitation to probe PRAS40?CmTOR interactions. They are also suited for cell proliferation and apoptosis assays, autophagy flux measurements (LC3-II turnover), and phenotypic screening of small-molecule inhibitors. The loss of AKT1S1 function is particularly valuable for drug discovery programs targeting the PI3K?CAKT?CmTOR axis, metabolic disease modeling, and autophagy research. For additional information or technical support, please contact Ascent Research.
