Interleukin-1\u03b2 drives NEDD8 nuclear-to-cytoplasmic translocation, fostering parkin activation via NEDD8 binding to the P-ubiquitin activating site

Meenakshisundaram Balasubramaniam,Martin R Farlow,Steven W Barger,Ling Liu,Chhanda Bose,W Sue T Griffin,Robert E Mrak,Richard A Jones,Paul A Parcon

doi:10.1186/s12974-019-1669-z

Abstract

BackgroundNeuroinflammation, typified by elevated levels of interleukin-1 (IL-1) α and β, and deficits in proteostasis, characterized by accumulation of polyubiquitinated proteins and other aggregates, are associated with neurodegenerative disease independently and through interactions of the two phenomena. We investigated the influence of IL-1β on ubiquitination via its impact on activation of the E3 ligase parkin by either phosphorylated ubiquitin (P-Ub) or NEDD8.MethodsImmunohistochemistry and Proximity Ligation Assay were used to assess colocalization of parkin with P-tau or NEDD8 in hippocampus from Alzheimer patients (AD) and controls. IL-1β effects on PINK1, P-Ub, parkin, P-parkin, and GSK3β—as well as phosphorylation of parkin by GSK3β—were assessed in cell cultures by western immunoblot, using two inhibitors and siRNA knockdown to suppress GSK3β. Computer modeling characterized the binding and the effects of P-Ub and NEDD8 on parkin. IL-1α, IL-1β, and parkin gene expression was assessed by RT-PCR in brains of 2- and 17-month-old PD-APP mice and wild-type littermates.ResultsIL-1α, IL-1β, and parkin mRNA levels were higher in PD-APP mice compared with wild-type littermates, and IL-1α-laden glia surrounded parkin- and P-tau-laden neurons in human AD. Such neurons showed a nuclear-to-cytoplasmic translocation of NEDD8 that was mimicked in IL-1β-treated primary neuronal cultures. These cultures also showed higher parkin levels and GSK3β-induced parkin phosphorylation; PINK1 levels were suppressed. In silico simulation predicted that binding of either P-Ub or NEDD8 at a singular position on parkin opens the UBL domain, exposing Ser65 for parkin activation.ConclusionsThe promotion of parkin- and NEDD8-mediated ubiquitination by IL-1β is consistent with an acute neuroprotective role. However, accumulations of P-tau and P-Ub and other elements of proteostasis, such as translocated NEDD8, in AD and in response to IL-1β suggest either over-stimulation or a proteostatic failure that may result from chronic IL-1β elevation, easily envisioned considering its early induction in Down’s syndrome and mild cognitive impairment. The findings further link autophagy and neuroinflammation, two important aspects of AD pathogenesis, which have previously been only loosely related.

Highlights

Neuroinflammation, with its associated elevation of interleukin-1 (IL-1) α and β synthesis and release, and autophagic failure, with its associated buildup of ubiquitinated protein aggregates in autophagosomes, are recognized as early events in the Alzheimer-related neuropathogenesis of Down’s syndrome [1,2,3,4] and, analoguously, of early events in the neuropathogenesis of Alzheimer’s disease (AD) [3, 5, 6]
Our molecular modeling predicts that activation of parkin by Neural precursor cell -expressed (NEDD8) is the result of NEDD8 binding to parkin at the same site that Ser65-phosphorylated ubiquitin (P-Ub) uses for its activation of parkin, suggesting that activation of parkin by either P-Ub or NEDD8 may serve as a necessary step in parkin-mediated ubiquitination of unwanted entities designated for lysosomal degradation
As a follow-up, in the PD-amyloid precursor protein (APP) mouse model of Alzheimer, we found that the synthesis of IL1α, IL-1β, and parkin was elevated in mice as early as 2 months of age (Fig. 1e, p = 0.036 for IL-1α, p = 0.04 for IL-1β, and p = 0.030 for parkin), months before Alzheimer lesions have been noted in this model [49]

Summary

Introduction

Neuroinflammation, with its associated elevation of interleukin-1 (IL-1) α and β synthesis and release, and autophagic failure, with its associated buildup of ubiquitinated protein aggregates in autophagosomes, are recognized as early events in the Alzheimer-related neuropathogenesis of Down’s syndrome [1,2,3,4] and, analoguously, of early events in the neuropathogenesis of Alzheimer’s disease (AD) [3, 5, 6] Such early events culminate in the neuropathological picture of end-stage AD, viz., the accumulation of neuronal and extraneuronal ubiquitinated protein aggregates. We investigated the influence of IL-1β on ubiquitination via its impact on activation of the E3 ligase parkin by either phosphorylated ubiquitin (P-Ub) or NEDD8

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Results

Discussion

Conclusion