The HTT Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal population targeting the HTT gene in human HAP1 cells. This product provides a diverse pool of gene-disrupted cells suitable for loss-of-function studies without clonal selection bias, ideal for pooled genetic screens and phenotypic assays.
HAP1 cells, derived from the KBM-7 chronic myeloid leukemia line, are a near-haploid human cell line widely used for genetic screening due to their stable karyotype and ease of manipulation. Originally a model for hematopoietic malignancies, HAP1 cells facilitate functional genomics and drug discovery applications.
The HTT gene encodes huntingtin, a large scaffold protein regulating vesicular trafficking, macroautophagy, and transcription. Huntingtin is cleaved by caspase-3 and calpain, phosphorylated by Akt and IKK??, and interacts with HIP1. It promotes autophagy by engaging ULK1, Beclin-1, ATG5, and LC3, and facilitates BDNF vesicle transport. In the nucleus, huntingtin binds the REST/NRSF complex to enable BDNF transcription. Consequently, HTT disruption impairs autophagic flux, reduces BDNF signaling, and alters mitochondrial dynamics via DRP1, compromising cellular stress responses.
In HAP1 cells, HTT knockout establishes an isogenic model for Huntington??s disease (HD) research. Despite their hematopoietic origin, these cells recapitulate HD-associated defects such as impaired autophagy, mitochondrial dysfunction, and calcium dysregulation. The polyclonal format allows robust assessment of bulk population phenotypes, enabling investigation of huntingtin??s interactions with optineurin and PGC-1?? and its role in coordinating mitochondrial quality control. This model is also valuable for studying pathways underlying polyglutamine toxicity and for validating therapeutic targets.
Researchers can utilize these cells in a variety of assays, including Western blotting and RT-qPCR for HTT expression analysis, immunofluorescence to assess huntingtin localization, and flow cytometry to measure apoptosis. Functional autophagy flux can be monitored via LC3 turnover and p62 accumulation, while mitochondrial function is evaluated by MTT assays and membrane potential dyes. Calcium imaging further delineates ER?Cmitochondrial connectivity. Additionally, the polyclonal cells serve as a clean background for expressing mutant HTT constructs to examine toxic gain-of-function, or for conducting compound screens to identify modulators of HD phenotypes. For detailed protocols and ordering, contact Ascent Research.
