Dominant Steady State Proteome Changes in the Absence of CHIP Highlight a Role in Neuronal Cell Membrane Integrity Linked to the Actin Cytoskeleton

Abstract

Functional integrity of the E3-ubiquitin ligase CHIP is essential for healthy aging in the brain. By virtue of its association with the core molecular chaperone machinery, studies into the physiological role(s) of CHIP have been dominated by its involvement in the quality control (QC) and degradation of unfolded, aggregated and/or mutated proteins. To elucidate the dominant changes in protein steady state levels in human neurons where CHIP function was ablated, we conducted an unbiased proteomic analysis in genetically engineered CHIP knockout compared to wild-type iPSC-derived cortical neurons. Rather than a broad effect on protein homeostasis, loss of CHIP function impacted on a focused cohort of proteins that analysis defined as actin cytoskeleton signalling and membrane integrity pathways. In support of the pathway analysis, CHIP KO cell-lines had enhanced sensitivity to mechanical and chemical membrane damage. Consistent with recent studies on the identification of a non-canonical HSP-independent E3-activity for CHIP, the lack of general QC defects in knock-out cells suggests that the major readout of CHIP function in cortical neurons is the regulation of native folded substrates involved in maintaining cellular ‘health’. Thus, CHIP-mediated regulation of the of cytoskeletal- and membrane-related proteins is likely to make a vital contribution to the neuroprotective activity of CHIP.