Physiological Mouse Brain Aβ Levels Are Not Related to the Phosphorylation State of Threonine-668 of Alzheimer's APP

Yoshitake Sano,Tadashi Nakaya,Steve Pedrini,Koichi Iijima,Kanae Iijima-Ando,Paul M Mathews,Shizu Takeda,Sam Gandy,Shigeyoshi Itohara,Toshiharu Suzuki

doi:10.1371/journal.pone.0000051

Abstract

BackgroundAmyloid-β peptide species ending at positions 40 and 42 (Aβ40, Αβ42) are generated by the proteolytic processing of the Alzheimer's amyloid precursor protein (APP). Aβ peptides accumulate in the brain early in the course of Alzheimer's disease (AD), especially Aβ42. The cytoplasmic domain of APP regulates intracellular trafficking and metabolism of APP and its carboxyl-terminal fragments (CTFα, CTFβ). The role of protein phosphorylation in general, and that of the phosphorylation state of APP at threonine-668 (Thr668) in particular, has been investigated in detail by several laboratories (including our own). Some investigators have recently proposed that the phosphorylation state of Thr668 plays a pivotal role in governing brain Aβ levels, prompting the current study.MethodologyIn order to evaluate whether the phosphorylation state of Thr668 controlled brain Aβ levels, we studied the levels and subcellular distributions of holoAPP, sAPPα, sAPPβ, CTFα, CTFβ, Aβ40 and Aβ42 in brains from “knock-in” mice in which a non-phosphorylatable alanyl residue had been substituted at position 668, replacing the threonyl residue present in the wild-type protein.ConclusionsThe levels and subcellular distributions of holoAPP, sAPPα, sAPPβ, CTFα, CTFβ, Aβ40 and Aβ42 in the brains of Thr668Ala mutant mice were identical to those observed in wild-type mice. These results indicate that, despite speculation to the contrary, the phosphorylation state of APP at Thr668 does not play an obvious role in governing the physiological levels of brain Aβ40 or Αβ42 in vivo.

Highlights

Alzheimer’s disease (AD) is characterized by abnormalities in posttranslational processing of two proteins, the amyloid precursor protein (APP) and the microtubule associated protein tau [1]
We have proposed a model wherein the phosphorylation state of APP at Thr668 may govern the state of activation of an intracellular signaling cascade across APP that leads to generation and translocation of APP intracellular domain fragment (AICD), analogous to the well-characterized signaling cascade across Notch that leads to generation and translocation of the Notch intracellular domain [15]
The potential importance of APP Thr668 phosphorylation was further emphasized in a recent review [3]. Because all these reports and reviews hinged on attribution of some biological significance to the phosphorylation state of APP Thr668, we investigated in vivo brain APP metabolism and Ab levels in mutant mice generated by knocking into their genome an APP gene containing a non-phosphorylatable alanyl substitution at position 668

Summary

Introduction

Alzheimer’s disease (AD) is characterized by abnormalities in posttranslational processing of two proteins, the amyloid precursor protein (APP) and the microtubule associated protein tau [1]. In order to evaluate whether the phosphorylation state of Thr668 controlled brain Ab levels, we studied the levels and subcellular distributions of holoAPP, sAPPa, sAPPb, CTFa, CTFb, Ab40 and Ab42 in brains from ‘‘knock-in’’ mice in which a non-phosphorylatable alanyl residue had been substituted at position 668, replacing the threonyl residue present in the wild-type protein. The levels and subcellular distributions of holoAPP, sAPPa, sAPPb, CTFa, CTFb, Ab40 and Ab42 in the brains of Thr668Ala mutant mice were identical to those observed in wild-type mice These results indicate that, despite speculation to the contrary, the phosphorylation state of APP at Thr668 does not play an obvious role in governing the physiological levels of brain Ab40 or Ab42 in vivo

Methods

Results

Conclusion