Reticulons RTN3 and RTN4‐B/C interact with BACE1 and inhibit its ability to produce amyloid β‐protein

Abstract β‐Secretase β‐site APP cleaving enzyme 1 (BACE1), is a membrane‐bound aspartyl protease necessary for the generation of amyloid β‐protein (Aβ), which accumulates in the brains of individuals with Alzheimer's disease (AD). To gain insight into the mechanisms by which BACE1 activity is regulated, we used proteomic methods to search for BACE1‐interacting proteins in human neuroblastoma SH‐SY5Y cells, which overexpress BACE1. We identified reticulon 4‐B (RTN4‐B; Nogo‐B) as a BACE1‐associated membrane protein. Co‐immunoprecipitation experiments confirmed a physical association between BACE1 and RTN4‐B, RTN4‐C (the shortest isoform of RTN‐4), and their homologue reticulon 3 (RTN3), both in SH‐SY5Y cells and in transfected human embryonic kidney (HEK) 293 cells. Overexpression of these reticulons (RTNs) resulted in a 30–50% reduction in the secretion of both Aβ40 and Aβ42 from HEK293 cells expressing the AD‐associated Swedish mutant amyloid precursor protein (APP), but did not affect Aβ secretion from cells expressing the APP β‐C‐terminal fragment (β‐CTF), indicating that these RTNs can inhibit BACE1 activity. Furthermore, a BACE1 mutant lacking most of the N‐terminal ectodomain also interacted with these RTNs, suggesting that the transmembrane region of BACE1 is critical for the interaction. We also observed a similar interaction between these RTNs and the BACE1 homologue BACE2. Because RTN3 and RTN4‐B/C are substantially expressed in neural tissues, our findings suggest that they play important roles in the regulation of BACE1 function and Aβ production in the brain.