Inefficient quality control of ribosome stalling during APP synthesis generates CAT-tailed species that precipitate hallmarks of Alzheimer\u2019s disease

Suman Rimal,Ishaq Tantray,Shuangxi Li,Yu Li,Charles Glabe,William W Seeley,Bingwei Lu,Su Guo,Rasika Vartak,Lea T Grinberg,Ji Geng,Salvatore Spina,Sungun Huh,Hannes Vogel

doi:10.1186/s40478-021-01268-6

Abstract

Amyloid precursor protein (APP) metabolism is central to Alzheimer’s disease (AD) pathogenesis, but the key etiological driver remains elusive. Recent failures of clinical trials targeting amyloid-β (Aβ) peptides, the proteolytic fragments of amyloid precursor protein (APP) that are the main component of amyloid plaques, suggest that the proteostasis-disrupting, key pathogenic species remain to be identified. Previous studies suggest that APP C-terminal fragment (APP.C99) can cause disease in an Aβ-independent manner. The mechanism of APP.C99 pathogenesis is incompletely understood. We used Drosophila models expressing APP.C99 with the native ER-targeting signal of human APP, expressing full-length human APP only, or co-expressing full-length human APP and β-secretase (BACE), to investigate mechanisms of APP.C99 pathogenesis. Key findings are validated in mammalian cell culture models, mouse 5xFAD model, and postmortem AD patient brain materials. We find that ribosomes stall at the ER membrane during co-translational translocation of APP.C99, activating ribosome-associated quality control (RQC) to resolve ribosome collision and stalled translation. Stalled APP.C99 species with C-terminal extensions (CAT-tails) resulting from inadequate RQC are prone to aggregation, causing endolysosomal and autophagy defects and seeding the aggregation of amyloid β peptides, the main component of amyloid plaques. Genetically removing stalled and CAT-tailed APP.C99 rescued proteostasis failure, endolysosomal/autophagy dysfunction, neuromuscular degeneration, and cognitive deficits in AD models. Our finding of RQC factor deposition at the core of amyloid plaques from AD brains further supports the central role of defective RQC of ribosome collision and stalled translation in AD pathogenesis. These findings demonstrate that amyloid plaque formation is the consequence and manifestation of a deeper level proteostasis failure caused by inadequate RQC of translational stalling and the resultant aberrantly modified APP.C99 species, previously unrecognized etiological drivers of AD and newly discovered therapeutic targets.

Highlights

Proteostasis denotes a cellular state in which protein synthesis, folding, and degradation are maintained at a homeostatic state such that an intact yet dynamic proteome is preserved [1, 2]
As the aggregated ER structures tended to be in contact with mitochondria, and ER-mitochondria contacts are involved in Ca2+ transfer from ER to mitochondria [52], we used a mito-GCaMP reporter to monitor mito-Ca2+ and observed increased mito-Ca2+ in Mhc > amyloid precursor protein (APP).C99 fly muscle (Additional file 1: Fig. S1a), suggesting that APP.C99 affects ER-mitochondrial Ca2+ signaling
In 5xFAD mouse brain synaptosome fractions, we found increased levels of certain ribosome-associated quality control (RQC) factors, including ZNF598 and Rack1, another sensor of stalled ribosomes [24], suggesting the (See figure on page.) Fig. 4 The ribosome stall sensor ZNF598 critically regulates the translational quality control and toxicity of APP.C99. a Immunoblots showing co-IP between APP.C99 and ZNF598 and other ribosome-associated QC factors. b Translocon pull-down with concanavalin A (ConA) beads showing increased ZNF598 and Ltn1 association with Sec61 translocon in APP.C99 transfected cells. c Immunoblots showing the effect of ZNF598 RNAi or OE on the level of stalled APP.C99. d Immunoblots showing the effect of ZNF598 RNAi on the level of mOC78-positive aberrant APP.C99 species

Summary

Introduction

Proteostasis denotes a cellular state in which protein synthesis, folding, and degradation are maintained at a homeostatic state such that an intact yet dynamic proteome is preserved [1, 2]. Proteostasis failure manifested as formation of aberrant protein aggregates, including the amyloid plaques composed of the β-amyloid (Aβ) peptide in AD [4, 5], is a defining feature of age-related neurodegenerative diseases [6,7,8]. The physiological function of APP remains incompletely understood [9], Aβ is known to be derived from the trafficking and processing of APP through the amyloidogenic pathway [10,11,12], whereby APP is cleaved by β-secretase (BACE) to generate soluble APP beta protein (sAPPβ) and APP. The origin of proteostasis failure and protein aggregate formation in the more common sporadic cases, is still enigmatic. The molecular nature of the initial seeding-activities and cellular mechanisms governing their formation remain largely undefined

Methods

Results

Conclusion