Invention novelty: This tangible material is a plasmid vector expressing a NKCC1-EGFP fusion protein.
Value Proposition: NKCC1 (Na+-K+-Cl- cotransporter 1) regulates cell volume and accumulation of intracellular Cl- and has been shown to promote invasiveness of brain tumor cells by modulating cell adhesion and cell migration. This vector is a tool that can facilitate a better understanding of NKCC1 protein and help in the identification of interacting proteins in different pathways affecting various disorders and diseases. Further it can be used for developing novel therapeutic strategies targeting not only NKCC1 but also other important interacting proteins. Other advantages include:
- Plasmid vector expressing a human NKCC1 gene fused with an enhanced green fluorescent protein (EGFP)
- Useful for expressing and localizing NKCC1 in mammalian cells lines due to the EGFP tag
- Can be used to generate stable cell lines expressing human NKCC1 tagged with EGFP for drug discovery
- Can be used for efficacy tests of therapeutic molecules on NKCC1
- Can be used to generate human NKCC1 antibodies
Technical Details: Johns Hopkins researchers have developed a mammalian lentiviral plasmid vector expressing an EGFP tagged NKCC1 protein. The construction of this vector involved generating a cDNA using RNA isolated from human astroctyic cell lines as the template followed by PCR amplification and subsequent cloning of the amplified product into the pcDNA3-EGFP plasmid vector. This vector was used to show interaction of NKCC1 protein with cellular cytoskeleton proteins, sub-cellular localization and its effect on the invasiveness and aggressiveness of glioblastoma cells. This vector can be useful for basic scientific studies, as a source to generate antibodies targeting NKCC1 or developing therapeutic strategies targeting NKCC1.
Publication(s): Garzon-Muvdi T, Schiapparelli P, ap Rhys C, Guerrero-Cazares H, Smith C, et al. (2012) Regulation of Brain Tumor Dispersal by NKCC1 Through a Novel Role in Focal Adhesion Regulation. PLoS Biol 10(5): e1001320. doi:10.1371/journal.pbio.1001320
