Dynamic physiological α-synuclein S129 phosphorylation is driven by neuronal activity

Abstract

In Parkinson’s disease and other synucleinopathies, the elevation of α-synuclein phosphorylated at Serine129 (pS129) is a widely cited marker of pathology. However, the physiological role for pS129 has remained undefined. Here we use multiple approaches to show for the first time that pS129 functions as a physiological regulator of neuronal activity. Neuronal activity triggers a sustained increase of pS129 in cultured neurons (200% within 4 h). In accord, brain pS129 is elevated in environmentally enriched mice exhibiting enhanced long-term potentiation. Activity-dependent α-synuclein phosphorylation is S129-specific, reversible, confers no cytotoxicity, and accumulates at synapsin-containing presynaptic boutons. Mechanistically, our findings are consistent with a model in which neuronal stimulation enhances Plk2 kinase activity via a calcium/calcineurin pathway to counteract PP2A phosphatase activity for efficient phosphorylation of membrane-bound α-synuclein. Patch clamping of rat SNCA−/− neurons expressing exogenous wild-type or phospho-incompetent (S129A) α-synuclein suggests that pS129 fine-tunes the balance between excitatory and inhibitory neuronal currents. Consistently, our novel S129A knock-in (S129AKI) mice exhibit impaired hippocampal plasticity. The discovery of a key physiological function for pS129 has implications for understanding the role of α-synuclein in neurotransmission and adds nuance to the interpretation of pS129 as a synucleinopathy biomarker.

Synuclein phosphorylation: pathogenic or physiologic?

“…In the current study, Ramalingam et al show that neural activity increases pSer-129 ~3-fold in primary culture, with no change in total α-synuclein 3 . This induction requires action potentials and synaptic transmission, indicating that network activity is responsible.…”

“…Dettmer, Selkoe and colleagues now address the physiological role of a post-translational modification associated with the synuclein + Lewy pathology characteristic of PD 3 . Considered specific for PD, phosphorylation of α-synuclein on Ser-129 (pSer-129) 4 has been widely used to assess the magnitude and extent of degeneration in animal models as well as the human condition.…”

“…Finally, Ramalingam et al (2023) address the functional role of Ser-129 phosphorylation 3 , a tall order given the unclear role of α-synuclein in neurotransmission studied in KO mice 1 , 2 . Introducing WT and mutant α-synuclein that cannot undergo phosphorylation at Ser-129 into α-synuclein KO neurons, they find that relative to WT, the mutant reduces the amplitude of excitatory postsynaptic currents (EPSCs) and increases the amplitude of inhibitory postsynaptic currents (IPSCs), with no change in their frequency.…”

Dynamic reversibility of α‐synuclein serine‐129 phosphorylation is impaired in synucleinopathy models

“…We recently showed that in the presence of liposomes, αS becomes a better substrate for S129 phosphorylation by Plk2 in a cell‐free system (Ramalingam et al , 2023). This prompted us to explore the relationship, if any, between αS solubility and pS129 for fPD‐linked αS mutants, some of which are known to interfere with membrane interaction (Fig 1A).…”

“…It has been estimated that 90% of αS in Lewy lesions contains phospho‐serine‐129 (pS129), connecting this post‐translational modification to the pathology (Fujiwara et al , 2002). However, we recently demonstrated that pS129 is also of physiological relevance (Ramalingam et al , 2023). We discovered that (i) pS129 arises in response to synaptic activity, (ii) activity‐dependent pS129 is reversible and not associated with toxicity, (iii) activity‐dependent pS129 is catalyzed by Polo‐like kinase 2 (Plk2) downstream of calcineurin and counteracted by protein phosphatase 2A, (iv) αS incapable of phosphorylation (S129A) reduces the ratio of excitatory versus inhibitory spontaneous post‐synaptic currents, and (v) hippocampal plasticity is impaired in our novel S129A knock‐in mouse model.…”