Postendocytotic traffic of the galanin R1 receptor: a lysosomal signal motif on the cytoplasmic terminus.
The neuropeptide galanin R1 receptor (GalR1) was tagged at its C terminus with EGFP (GalR1-EGFP) to study receptor localization and trafficking. In PC12 and HEK293 cells, functional GalR1-EGFP was expressed on the plasma membrane and internalized into cytoplasmic vesicles after galanin stimulation. The internalization was blocked by 0.4 M sucrose and by silencing of clathrin with siRNA methodology. Internalized GalR1-EGFP and LysoTracker, a lysosomal marker, overlapped in intracellular vesicles after prolonged galanin stimulation. This colocalization was strongly reduced after site-directed mutagenesis of the motif YXXØ on the C terminus of GalR1 (where Ø is a bulky hydrophobic residue and X any amino acid). Taken together, these data suggest that GalR1 is internalized via the clathrin-dependent, endocytic pathway and then, to a large extent, delivered to lysosomes for degradation through the lysosome-targeting signal YXXØ.
Xia
Sheng
S
Departments of Neuroscience and Physiology and Pharmacology, Karolinska Institutet, S-171 77 Stockholm, Sweden.
Dun
Xing-Peng
XP
Hu
Ping-Sheng
PS
Kjaer
Svend
S
Zheng
Kang
K
Qian
Yu
Y
Solén
Christina
C
Xu
Tao
T
Fredholm
Bertil
B
Hökfelt
Tomas
T
Xu
Zhi-Qing David
ZQ
D020816:Amino Acid Motifs / Q000502:physiology; D002460:Cell Line; D002966:Clathrin; D003593:Cytoplasm / Q000378:metabolism; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism; D019004:Galanin / Q000494:pharmacology; D049452:Green Fluorescent Proteins; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D021381:Protein Transport; D044091:Receptor, Galanin, Type 1 / Q000378:metabolism*; D013395:Sucrose
D002966:Clathrin; D044091:Receptor, Galanin, Type 1; D049452:Green Fluorescent Proteins; D013395:Sucrose; D019004:Galanin