Neto1 associates with the NMDA receptor/amyloid precursor protein complex.

Neuropilin tolloid-like 1 (Neto1), is a CUB domain-containing transmembrane protein that was recently identified as a novel component of the NMDA receptor complex. Here, we have investigated the possible association of Neto1 with the amyloid precursor protein ( APP)695/GluN1/GluN2A and APP695/GluN1/GluN2B NMDA receptor trafficking complexes that we have previously identified. Neto1 ^HA was shown to co-immunoprecipitate with assembled NMDA receptors via GluN2A or GluN2B subunits; Neto1 ^HA did not co-immunoprecipitate APP695 ^FLAG. Co-immunoprecipitations from mammalian cells co-transfected with APP695 ^FLAG, Neto1 ^HA and GluN1/GluN2A or GluN1/GluN2B revealed that all four proteins co-exist within one macromolecular complex. Immunoprecipitations from native brain tissue similarly revealed the existence of a GluN1/GluN2A or GluN2B/ APP/Neto1 complex. Neto1 ^HA caused a reduction in the surface expression of both NMDA receptor subtypes, but had no effect on APP695 ^FLAG- or PSD-95α^c-Myc enhanced surface receptor expression. The Neto1 binding domain of GluN2A was mapped using GluN1/GluN2A chimeras and GluN2A truncation constructs. The extracellular GluN2A domain does not contribute to association with Neto1 ^HA but deletion of the intracellular tail resulted in a loss of Neto-1 ^HA co-immunoprecipitation which was paralleled by a loss of association between GluN2A and SAP102. Thus, Neto1 is concluded to be a component of APP/ NMDA receptor trafficking complexes. [ABSTRACT FROM AUTHOR]