Growth Factor Receptor-Bound Protein 2 Promotes Autophagic Removal of Amyloid-β Protein Precursor Intracellular Domain Overload in Neuronal Cells

Abstract

The ascertainment of elevated levels of amyloid-β protein precursor intracellular domain (AICD) in Alzheimer's disease (AD) brains and the fact that it contributes to AD-like pathology has geared the search toward a new paradigm. While studying endogenous as well as overexpressed Grb2-AICD interaction in AD cell models, it was found that Grb2 co-localized to compartments along with AICD. We report now that these vesicles form in a clathrin and dynamin independent manner. Both types of vesicles mature into autophagosomes, merge with lysosomes, and relieve the cells of AICD overload. Inhibiting autophagosome formation results in vesicle accumulation. AICD-level is reduced in Grb2 excess condition in Cycloheximide Chase setup. Reduced caspase activity and apoptosis point toward the fact that the cytotoxic effect of AICD is alleviated by its sequestration in autolysosomes. Hence we state that the entrapping of AICD in Grb2 vesicles and its clearance via autophagosomes is a survival contrivance on the part of the cell. This study unravels, for the first time, the roles of Grb2 in autophagy and in handling toxic protein overload in an AD-like scenario.

Cellular levels of growth factor receptor bound protein 2 (Grb2) and cytoskeleton stability are correlated in a neurodegenerative scenario

“…Given that Grb2 is naturally overexpressed in whole brain lysates of AD patients and also that it sequesters AICD in vesicles (McShea et al, 1999; Raychaudhuri and Mukhopadhyay, 2010; Roy et al, 2014), it was reasonable to hypothesize that Grb2 might reverse the cytoskeletal protein degradation in AD-like conditions. Restoration in both protein and transcript levels of the cytoskeleton proteins were investigated upon Grb2 overexpression [Fig.…”

“…Several reports on Grb2 strongly suggest its role in survival in AD through sequestration of AICD via autophagy (Raychaudhuri and Mukhopadhyay, 2010; Roy et al, 2014). The present study for the first time demonstrates that Grb2 is naturally overexpressed in AD whole human brain lysate and also in the APP/PS1 AD mouse model [Fig.…”

“…A similar increase in RhoA has been reported in SHSY-5Y cells by Petratos et al (2008), supporting our assumption that the simultaneous abundance of Aβ, AICD and Grb2 in and around these cells could be a true reflection of an AD-like condition. Elevation of Cdc42 activity is justified as it is known to be agonistically regulated with autophagy by phosphatidylethanolamine-conjugated LC3 proteins (LC3II) (Chung et al, 2012), and our group has previously demonstrated autophagy induction in a similar context (Roy et al, 2014). Considering that Rac1 activity decreases significantly in our model, the expression of PAK1, the downstream effector of both Cdc42 and Rac1, relies on the fine balance of the activities of both.…”

“…Recently, our group has shown that excess AICD under AD-like conditions is sequestered from the system by Grb2 through the autophagic pathway (Raychaudhuri and Mukhopadhyay, 2010; Roy et al, 2014). In general, Grb2, a cytoplasmic protein, plays a key role in Ras-mediated growth factor signalling, proliferation and the cell cycle.…”

Ubiquitin mediated regulation of the E3 ligase GP78 by Mahogunin in trans affects mitochondrial homeostasis

“…Immortal melanocytes were grown in culture as described previously (Srivastava and Chakrabarti, 2014;Hida et al, 2009). SH-SY5Y cells (a gift from Debashis Mukhopadhyay) and MEF cells (CF-1 strain; a gift from Mitradas M. Panicker, National Centre for Biological Sciences, Tata Institute of Fundamental Research, Bangalore, India) were also grown under standard cell culture conditions as described previously (Srivastava and Chakrabarti, 2014;Roy et al, 2013). Immortal melanocytes (a gift from Ramanujan S. Hegde) were obtained from the Wellcome Trust Functional Genomics Cell Bank.…”