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

Abstract

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.

• Authors: Ana Jorge-Finnigan, Rune Kleppe, Kunwar Jung-Kc, Ming Ying, Michael Marie, Ivan Rios-Mondragon, Michael F Salvatore, Jaakko Saraste, Aurora Martinez
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 292 Issue 34 Pg. 14092-14107 (08 25 2017) ISSN: 1083-351X [Electronic] United States
• PMID: 28637871 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Chemical References

• Nerve Tissue Proteins
• Recombinant Fusion Proteins
• Green Fluorescent Proteins
• Serine
• Tyrosine 3-Monooxygenase

Topics

• Amino Acid Substitution
• Animals
• Cell Line, Tumor
• Dopaminergic Neurons (cytology, enzymology, metabolism)
• Golgi Apparatus (enzymology, metabolism)
• Green Fluorescent Proteins (genetics, metabolism)
• HEK293 Cells
• Humans
• Microscopy, Confocal
• Microscopy, Fluorescence
• Microtubules (enzymology, metabolism)
• Mutagenesis, Site-Directed
• Mutation
• Nerve Tissue Proteins (genetics, metabolism)
• Phosphorylation
• Protein Processing, Post-Translational
• Protein Transport
• Rats
• Recombinant Fusion Proteins (metabolism)
• Serine (metabolism)
• Synaptic Vesicles (enzymology, metabolism)
• Tyrosine 3-Monooxygenase (genetics, metabolism)