Posttranslational Modifications of α-Synuclein, Their Therapeutic Potential, and Crosstalk in Health and Neurodegenerative Diseases

Kambiz Hassanzadeh; Jun Liu; Santhosh Maddila; M. Maral Mouradian

doi:10.1124/pharmrev.123.001111

Article Figures & Data

Figures

Tables

Download figure
Open in new tab
Download powerpoint
Download figure
Open in new tab
Download powerpoint
Fig. 1
α-Synuclein posttranslational modifications. Summary of the various posttranslational modifications of α-Syn categorized by the type of modification including addition of chemical groups (red), a polypeptide (green), complex molecules (black), and modifications involving amino acids (blue). 4-Hydroxynonenal is another reported posttranslational modification not depicted in this figure.
Download figure
Open in new tab
Download powerpoint
Fig. 2
α-Synuclein structure with posttranslational modification sites. Created with BioRender.com pursuant to its Academic License Terms.
Download figure
Open in new tab
Download powerpoint
Fig. 3
Schematic diagrams of the impact of posttranslational modifications on α-Syn. The three domains of α-Syn are depicted: the N-terminal domain (1–60), the central region (61–95), and the C-terminal domain (96–140). PTMs can a) promote (red circles) or inhibit (green circles) α-Syn aggregation, b) promote (red circles) or inhibit (green circles) toxicity, and c) inhibit (red) or promote (green) clearance. Polyamination that targets acidic and negatively charged residues promotes α-Syn aggregation (not shown in the diagram). Created with BioRender.com pursuant to its Academic License Terms.
Download figure
Open in new tab
Download powerpoint
Fig. 4
Crosstalk among posttranslational modifications of α-synuclein. An illustration of the complex interplay of α-Syn PTMs highlighting the dynamic relationship between various PTMs. Given that phosphorylation is the most studied PTM of α-Syn, the majority of research on PTMs crosstalk focuses on interactions between phosphorylation and other PTMs. a) Phosphorylation and ubiquitination: Increased phosphorylation of α-Syn at S129 is associated with enhanced ubiquitination at all lysine residues (K-Ub: indicated by a bracket), whereas di-ubiquitination or tetra-ubiquitination at K12 inhibits the phosphorylation of α-Syn at S129. b) Phosphorylation and SUMOylation: The impact of SUMOylation on α-Syn phosphorylation at S129 is kinase-dependent. SUMOylation of α-Syn prevents PLK2-mediated phosphorylation at S129. However, G protein-coupled receptor kinase 5 mediated phosphorylation at S129 is not impacted by the cellular SUMO machinery. c) Phosphorylation and O-GlcNAcylation: O-GlcNAcylation at T72 slightly enhances phosphorylation at S87 by CK1 but prevents phosphorylation at S129 by all three kinases. d) Phosphorylation and acetylation: Although there is presently no empirical evidence substantiating a direct correlation between the phosphorylation and acetylation of α-Syn, studies employing pharmacological methods have shown a negative association between these two PTMs, suggesting that N-terminal acetylation or lysine acetylation (indicated by a bracket) could potentially impede α-Syn phosphorylation. e) Phosphorylation and nitration: the C-terminal tyrosine residues (Y125, Y133, and Y136) of α-Syn are targets for phosphorylation and nitration. Specifically, nitration at Y133 inhibits phosphorylation at S129. f) Ubiquitination and SUMOylation: ubiquitination and SUMOylation may engage in cooperative or competitive interactions since both these PTMs aim to modify the lysine residues of α-Syn. g) Ubiquitination and glycation: the N-terminal lysine residues of α-Syn are potential targets for both glycation and ubiquitination. Evidence indicates that α-Syn glycation inhibits its ubiquitination. K-Ac, acetylation at lysine residues; K-Ub, ubiquitination at lysine residues; N,nitration; N-Ac, N terminal acetylation; O-GN, O-GlcNAcylation; P, phosphorylation; Ub, ubiquitination. Created with BioRender.com pursuant to its Academic License Terms.

Tables

Figures

View popup

TABLE 1

α-Synuclein posttranslational modifications

PTM	Modifying Group	Enzymes Involved	Site and Amino Acid Involved	Functional Effects
Phosphorylation	Phosphate (PO4-3)	Casein kinases (CK1, CK2) (Takahashi et al., 2007; Waxman and Giasson, 2008) Glycogen synthase kinase-3β (Hu et al., 2020; Takaichi et al., 2020) Polo like kinase 2 (Inglis et al., 2009) Death-associated protein kinase 1 (Shin and Chung, 2020) Inflammation-associated serine-threonine kinase, PKR (EIF2AK2) (Reimer et al., 2018) G protein-coupled receptor kinases including GRK2 (Pronin et al., 2000), GRK3 (Sakamoto et al., 2009), GRK5 (Pronin et al., 2000; Arawaka et al., 2006), and GRK6 (Sakamoto et al., 2009) Protein phosphatase 2 A (Lee et al., 2011)	Serine S87, S129 (Chen et al., 2009a; Xu et al., 2015), S42 (Zhang et al., 2023)	S129: Promotes aggregation and some data reported decreases aggregation S87: Inhibits aggregation
			Tyrosine Y39, Y125, Y133, Y136 (Okochi et al., 2000; Kleinknecht et al., 2016; Manzanza et al., 2021)	Y39: Promotes aggregation Y125: Inhibits aggregation Y133: Protective, decreases in the seeding potency Y136: Inhibits aggregation
			Threonine T64, T72, T75 and T81 (Zhang et al., 2023)	T64: Promotes oligomerization
Ubiquitination	Ubiquitin a small (8.6 kDa) protein	Activating (E1), conjugating (E2), and ligating (E3) enzymes (Hershko and Ciechanover, 1998) E3 ubiquitin ligase SIAH (seven in absentia homolog) (Liani et al., 2004; Lee et al., 2008) Nedd4 ubiquitin ligases (Tofaris et al., 2011; Davies et al., 2014; Sugeno et al., 2014; Wijayanti et al., 2015) Protein de-ubiquitinase enzymes: USP13 (Moussa, 2016), USP9X (Rott et al., 2011), USP8 (Alexopoulou et al., 2016)	Lysine residues (Tofaris et al., 2003; Anderson et al., 2006)	Promotes degradation
SUMOylation	SUMO proteins (SUMO1-3)	SUMOylation enzymes: activating (E1), conjugating (E2: Ubc9), and ligating (E3) enzymes (Geiss-Friedlander and Melchior, 2007) DeSUMOylation enzyme: sentrin/small ubiquitin-like modifier-specific protease (Wilkinson and Henley, 2010)	Lysine residues: mostly K96 and K102 (Dorval and Fraser, 2006; Krumova et al., 2011; Rott et al., 2017)	Debatable: promotes or inhibits aggregation
Acetylation	Acetyl group (CH3CO)	N-terminal acetyltransferases (Deng et al., 2020). These enzymes include NatA, NatB, NatC, NatD, NatE, NatF, and NatH (Aksnes et al., 2019)	Lysine acetylation (Nε-acetylation) and N-terminal protein acetylation (Nα-acetylation) (Kang et al., 2012, 2013; Lundby et al., 2012; Bartels et al., 2014; Dikiy and Eliezer, 2014; Bu et al., 2017; Ruzafa et al., 2017; Deng et al., 2020; Runfola et al., 2020; Vinueza-Gavilanes et al., 2020)	N-terminal acetylation reduces α-Syn oligomerization
Glycosylation	N-acetylglucosamine (GlcNAc)	O-GlcNAc transferase and O-GlcNAcase (Wani et al., 2017)	O-glycosylation: Serine 87 and Threonine 72 (Alfaro et al., 2012; Marotta et al., 2015; Zhang et al., 2017a; Lewis et al., 2017) Threonine 54, 64 (Alfaro et al., 2012), Threonine 54 and 75 (Zhang et al., 2023)	T72: Inhibits aggregation S87: Inhibits aggregation T75: Prevents the extension of PFFs T81: Prevents the extension of PFFs
Glycation	Glucose, fructose, and their derivatives	Nonenzymatic	Lysine residues mostly at K6, K10, K12, K21, K23, K32, K34, and K43 and K45 (Vicente Miranda et al., 2017b)	Promotes oligomerization and aggregation
	Carboxymethyl group (-CH2-COOH)	Nonenzymatic	Lysine residues: K12, K21, K23, K32, K34, K45, K58, K60, K80, K96, K102 (Zhang et al., 2023)
	Carboxyethyl group (-CH2-CH2-COOH)	Nonenzymatic	Lysine residues: K12, K60 (Zhang et al., 2023)
Nitration	Nitric oxide	Nonenzymatic	Tyrosine residue: Y39, Y125, Y133, and Y136 (Giasson et al., 2000; Norris et al., 2003; Yamin et al., 2003; Hodara et al., 2004; Uversky et al., 2005; Danielson et al., 2009; Sevcsik et al., 2011)	Y39: Promotes oligomerization Y125: Promotes dimerization Y133: Promotes aggregation Y136: Promotes aggregation
Oxidation	Reactive oxygen species	Nonenzymatic	Methionine residue: M1, M5, M116, and M127 (Uversky et al., 2002; Hokenson et al., 2004; Zhou et al., 2010; Chavarría and Souza, 2013; Maltsev et al., 2013; Schildknecht et al., 2013; Ponzini et al., 2019)	Inhibits fibrillization
Polyamination	Polyamines, small cationic molecules	Transglutaminase (Folk et al., 1980) Polyamine catabolism enzymes: SMOX and SAT1 (Lewandowski et al., 2010; Zahedi et al., 2020)	Acidic and negatively charged residues (Antony et al., 2003; Fernández et al., 2004)	Promotes aggregation
Arginylation	Arginine	Arginyltransferase (ATE1) (Saha and Kashina, 2011)	Glutamate residue: E46 and E83 (Waxman et al., 2010; Wang et al., 2017a; Boyer et al., 2020; Pan et al., 2020), and E57 (Zhang et al., 2023)	E83: Reduces aggregation E46: Slows down the fibrillization
Truncation	No modifying group	Neurosin (Kasai et al., 2008), calpain I (Dufty et al., 2007), cathepsin D (Sevlever et al., 2008), and matrix metalloproteinase 3 (Choi et al., 2011).	No specific residue	C-terminus: Promotes aggregation NAC region: inhibit aggregation N-terminal: Slows down the fibrillization but increases toxicity
Methylation Dimethylation Trimethylation	Methyl group (-CH3)	Methyltransferase	Methylation: Lysine residues: K12, K21, K23, K34, K45, K58, K60, K80, K96 Dimethylation: Lysine residues: K12, K21, K58, K60, K96, K102 Trimethylation: Lysine residue: K60 (Zhang et al., 2023)	No data
4-hydroxynonenal	4-hydroxynonenal group	Nonenzymatic	Lysine residue: K60 (Zhang et al., 2023)	No data

View popup

TABLE 2

Pharmacological tools targeting α-synuclein PTMs

PTM	Pharmacological Intervention	Type of study and model	Effect on PTM	Outcome
Phosphorylation	Eicosanoyl-5-hydroxytryptamide, a fatty acid derivative of serotonin found in coffee	Thy1-Syn transgenic mice	Reduces α-Syn phosphorylation	Reduces α-Syn aggregation in the brain (Lee et al., 2011)
	Eicosanoyl-5-hydroxytryptamide	The α-Syn PFF inoculation model in C57BL/6 J mice	Reduces the accumulation of phosphorylated α-Syn	Reduces the spread of α-Syn PFF and neuroinflammation and improves behavioral performance (Yan et al., 2018)
	BI 2536, small-molecule PLK inhibitor	Syn-GFP mice	Reduces phosphorylation of α-syn at S129	No effect on α-Syn aggregation (Weston et al., 2021b)
Ubiquitination	BK50118-C, a small molecule inhibitor of ubiquitin-specific protease-13	Lentiviral vector expressing human α-Syn injected in the substantia nigra of mice	Increases ubiquitination and proteasome activity	Elevates dopamine levels and enhances motor skills (Liu et al., 2022)
Ubiquitination	Canthin-6-one	PC12 cells	Upregulation of the PSMD1 gene and activating the UPS	α-syn degradation (Yuan et al., 2019)
SUMOylation	Ginkgolic acid, the SUMO inhibitor	Rat cortical primary neurons. SH-SY5Y neuroblastoma cells.	Prevents the aggregation of α-Syn while boosting the presence of LC3-positive autophagosomes within cells	Promotes macroautophagy and removes aggregates (Vijayakumaran et al., 2019)
Glycosylation	ASN90/ASN120290/ ASN-561, a glycoside hydrolase O-GlcNAcase inhibitor	Thy1-Syn transgenic mice	Increases O-GlcNAcylated α-Syn	Improves motor skills and reduces astrogliosis (Permanne et al., 2022)
Glycosylation	Thiamet-G, a selective inhibitor of the enzyme O-GlcNAcase	Cell types, including mouse primary cortical neurons	Increases O-GlcNAcylated α-Syn	Reduces the uptake of α-Syn fibrils by cells (Tavassoly et al., 2021)
Glycation	The dicarbonyl compound MGO	Thy1-Syn transgenic mice	Induces α-Syn glycation	Induces the formation of α-synuclein aggregates and disrupts synaptic transmission (Vicente Miranda et al., 2017b)
Glycation	Aminoguanidine and tenilsetam: MGO scavengers	H4 cells and Drosophila	Decreases MGO levels and inhibit glycation	Attenuate α-synuclein aggregation and toxicity and improve the motor performance of α-synuclein expressing flies (Vicente Miranda et al., 2017b)
Nitration	MK801, NMDA receptor antagonist	Mice expressing human α-Syn on SNCA knockout background and neuron-glia cocultures	Reduces α-Syn nitration	Prevents LPS-induced DA neuronal death (Gao et al., 2008)
Nitration	GYY4137, an H2S slow-releasing compound	MPTP mouse model of PD	Reduces α-syn nitration	Exerts neuroprotection (Hou et al., 2017)
Polyamination	Cystamine and cysteamine, Transglutaminase inhibitors	Double knockout mice (Smox/Sat1-dKO): deletion of principal polyamine catabolic enzymes	Reduces polyamine expression	Reduces α-Syn expression and aggregation, the severity of cerebellar injury, and ataxia (Zahedi et al., 2020)
Polyamination	DENSPM (N1, N11-diethylnorspermine), a polyamine analog	Thy1-Syn transgenic mice	Increases SAT1 activity and reduces polyamines	Reduces α-Syn aggregation in the basal ganglia and improves the PD phenotype (Lewandowski et al., 2010)
Truncation	Calpeptin, a calpain inhibitor	MPTP-induced mouse model of PD	Reduces α-Syn aggregation	Attenuates gliosis and inflammatory markers and reduces α-Syn aggregation (Haque et al., 2020)

View popup

TABLE 3

Crosstalk between α-synuclein posttranslational modifications

PTM	Crosstalk with ….	Location	Crosstalk Type	Mechanism
Phosphorylation	O-GlcNAcylation	Reciprocal S87^a	Negative Intraprotein	O-GlcNAcylation at S87 may prevent the effect of phosphorylation on α-Syn membrane binding (Lewis et al., 2017)
		Proximal T72^b	Positive Intraprotein	O-GlcNAcylation at T72 promotes phosphorylation at S87 (Marotta et al., 2015)
		Distal T72^c	Negative Intraprotein	O-GlcNAcylation at T72 prevents phosphorylation at S129 (Marotta et al., 2015)
	SUMOylation	Distal Lysine residues	Positive Intraprotein	PTMs work together to regulate the α-Syn autophagic and proteasomal degradation (Shahpasandzadeh et al., 2014)
	SUMOylation	Distal Lysine residues	Negative Intraprotein	SUMOylation of α-Syn down-regulates phosphorylation at S129 (Shahpasandzadeh et al., 2014)
	Ubiquitination	Distal Lysine residues	Negative Intraprotein	Ubiquitination at K12 prevents α-Syn phosphorylation at S129 (Haj-Yahya et al., 2013)
	Ubiquitination	Distal Lysine residues	Positive Intraprotein	Higher phosphorylation of α-Syn results in greater ubiquitination (Shahpasandzadeh et al., 2014)
	Nitration	Proximal	Negative Intraprotein	Nitration at Y133 prevents the protective S129 phosphorylation (Kleinknecht et al., 2016)
Ubiquitination	SUMOylation	Reciprocal Lysine residues	Negative Intraprotein	Antagonistically one prohibits the other (Rott et al., 2017; Rousseaux et al., 2018; Savyon and Engelender, 2020)
	SUMOylation	Reciprocal Lysine residues	Positive Intraprotein	Impaired α-Syn SUMOylation inhibits the UPS (Zhu et al., 2018)
	Glycation	Reciprocal Lysine residues	Negative Intraprotein	Glycation blocks α-Syn ubiquitination (Vicente Miranda et al., 2017b)
Oxidation	Oxidation	Proximal and distal M1, M5,M49	Negative Intraprotein	M-oxidation in one site controls other Met oxidation, influencing the α-Syn membrane affinity (Maltsev et al., 2013)

↵^aReciprocal: crosstalk at the same residue.
↵^bProximal: crosstalk at nearby sites.
↵^cDistal: crosstalk at distal sites.