J. Cell Biol.
© The Rockefeller University Press
0021-9525/97/08/671/10 $2.00
Volume 138, Number 3, August 11, 1997 671-680

Novel -Secretase Cleavage of -Amyloid Precursor Protein in the Endoplasmic Reticulum/Intermediate Compartment of NT2N Cells

Abraham S.C. Chyung,^* Barry D. Greenberg, David G. Cook,^* Robert W. Doms,^* and Virginia M.-Y. Lee^*

^* Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104; and  Cephalon, Incorporated, West Chester, Pennsylvania 19380

Previous studies have demonstrated that NT2N neurons derived from a human embryonal carcinoma cell line (NT2) constitutively process the endogenous wild-type -amyloid precursor protein (APP) to amyloid  peptide in an intracellular compartment. These studies indicate that other proteolytic fragments generated by intracellular processing must also be present in these cells. Here we show that the NH₂-terminal fragment of APP generated by -secretase cleavage (APP) is indeed produced from the endogenous full length APP (APP_FL). Pulse-chase studies demonstrated a precursor-product relationship between APP_FL and APP as well as intracellular and secreted APP fragments. In addition, trypsin digestion of intact NT2N cells at 4°C did not abolish APP recovered from the cell lysates. Furthermore, the production of intracellular APP from wild-type APP appears to be a unique characteristic of postmitotic neurons, since intracellular APP was not detected in several non-neuronal cell lines. Significantly, production of APP occurred even when APP was retained in the ER/ intermediate compartment by inhibition with brefeldin A, incubation at 15°C, or by expression of exogenous APP bearing the dilysine ER retrieval motif.

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