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Novel Construct to Study the Functional Effects of SNPs on TrkA Protein Interactions and Localization

  • Author(s): Chin, Brian Wai Wen
  • et al.
Abstract

A unique family has previously been reported where medullary cystic kidney disease and bipolar disorder segregated very closely together (Kimmel et al, 2005). This lead to the isolation of a novel SNP (rs144901788) in the NTRK1 gene. NTRK1 codes for a neurotrophic receptor tyrosine kinase known as TrkA that signals down well-known survival and mitogenic pathways such as MAPK/ERK. Understanding how TrkA function is modified in these individuals may help shed light on a possible mechanism for bipolar disorder. In order to examine any functional effects said SNP may have TrkA function, we procured a construct from Kazusa containing TrkA with Promega's proprietary HaloTag® conjugated to the N-terminus. Based off the initial results with the construct, we determined that protein product would not provide an accurate reproduction of TrkA function. As a result, we produced an alternate construct capable of producing tagged, recombinant TrkA. Here, we characterize the products of said construct via western blot and live-cell fluorescent visualization and demonstrate its suitability as a model for studying SNPs in TrkA

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