The EIPR1 Knockout HEK293T Polyclonal Cells comprise a CRISPR/Cas9-edited polyclonal knockout cell population designed to disrupt the EIPR1 gene in the human embryonic kidney HEK293T cell line. This polyclonal pool provides a genetically heterogeneous loss-of-function model, enabling investigation of EIPR1-dependent cellular processes without clonal selection biases. The targeted gene disruption abolishes functional EIPR1 protein expression, making these cells a valuable tool for studying endosomal maturation and autophagy regulation.
HEK293T cells are a widely utilized human embryonic kidney epithelial cell line stably expressing the SV40 large T-antigen, which promotes episomal replication of plasmids containing the SV40 origin and enhances transient protein expression. These cells exhibit robust growth, high transfection efficiency, and are routinely employed for mechanistic studies, protein production, and functional genomics. Their epithelial origin and active endolysosomal system make them an appropriate host for dissecting EIPR1-mediated membrane trafficking events.
EIPR1 functions as a critical regulator of autophagy and endocytic trafficking by localizing to early endosomes where it interacts with RAB5 and EEA1. Upon autophagy induction, downstream of nutrient deprivation or mTORC1 inhibition, EIPR1 facilitates autophagosome-lysosome fusion by recruiting the HOPS tethering complex components VPS16 and VPS33A, along with the SNARE proteins STX17, SNAP29, and VAMP8. This recruitment drives SNARE-mediated membrane fusion, enabling autophagic cargo delivery to lysosomes and subsequent degradation. Consequently, EIPR1 loss impairs autophagic flux and endosome maturation.
In HEK293T cells, disruption of EIPR1 allows detailed examination of autophagy and endolysosomal pathway dynamics in a well-characterized, easily manipulated system. Although HEK293T cells are not derived from neural tissue, the conserved nature of the autophagic machinery permits mechanistic insights relevant to neurodevelopmental disorders associated with EIPR mutations, such as epilepsy, microcephaly, and intellectual disability. The polyclonal knockout pool ensures that phenotypes observed are not artifacts of single-cell cloning, providing a more representative model of gene disruption.
These EIPR1 knockout polyclonal cells support a wide range of experimental approaches, including Western blot analysis of LC3 lipidation to monitor autophagosome formation, autophagic flux measurements using lysosomal inhibitors, co-immunoprecipitation to study protein interactions with HOPS and SNARE components, and immunofluorescence-based endocytosis assays. Researchers can employ this model for drug screening aimed at identifying autophagy modulators or for dissecting molecular mechanisms underlying EIPR1-related pathologies. For additional information or to discuss custom applications, please contact Ascent Research.
