Neurobiology
Phosphorylation at serine 31 targets tyrosine hydroxylase to vesicles for transport along microtubules

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Tyrosine hydroxylase (TH) catalyzes the conversion of l-tyrosine into l-DOPA, which is the rate-limiting step in the synthesis of catecholamines, such as dopamine, in dopaminergergic neurons. Low dopamine levels and death of the dopaminergic neurons are hallmarks of Parkinson's disease (PD), where α-synuclein is also a key player. TH is highly regulated, notably by phosphorylation of several Ser/Thr residues in the N-terminal tail. However, the functional role of TH phosphorylation at the Ser-31 site (THSer(P)-31) remains unclear. Here, we report that THSer(P)-31 co-distributes with the Golgi complex and synaptic-like vesicles in rat and human dopaminergic cells. We also found that the TH microsomal fraction content decreases after inhibition of cyclin-dependent kinase 5 (Cdk5) and ERK1/2. The cellular distribution of an overexpressed phospho-null mutant, TH1-S31A, was restricted to the soma of neuroblastoma cells, with decreased association with the microsomal fraction, whereas a phospho-mimic mutant, TH1-S31E, was distributed throughout the soma and neurites. TH1-S31E associated with vesicular monoamine transporter 2 (VMAT2) and α-synuclein in neuroblastoma cells, and endogenous THSer(P)-31 was detected in VMAT2– and α-synuclein–immunoprecipitated mouse brain samples. Microtubule disruption or co-transfection with α-synuclein A53T, a PD-associated mutation, caused TH1-S31E accumulation in the cell soma. Our results indicate that Ser-31 phosphorylation may regulate TH subcellular localization by enabling its transport along microtubules, notably toward the projection terminals. These findings disclose a new mechanism of TH regulation by phosphorylation and reveal its interaction with key players in PD, opening up new research avenues for better understanding dopamine synthesis in physiological and pathological states.

confocal microscopy
dopamine
Parkinson disease
phosphorylation
vesicles
axonal transport
tyrosine hydroxylase phosphorylated at Ser-31
tyrosine hydroxylase

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This work was supported by Research Council of Norway Grant FRIMEDBIO 214012 and the K. G. Jebsen Foundation. The authors declare that they have no conflicts of interest with the contents of this article.

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The abbreviations used are: PD, Parkinson's disease; l-DOPA, l-3,4-dihydroxyphenylalanine; GC, Golgi complex; TH, tyrosine hydroxylase; DA, dopamine; VMAT2, vesicular monoamine transporter 2; NGF, neuronal growth factor; α-syn, α-synuclein; R/S, roscovitine and SL327; sytI, synaptotagmin I; PLA, proximity ligation assay; WGA, wheat germ agglutinin; CVM, chromaffin vesicle membrane.