Interaction of γ-COP with a transport motif in the D1 receptor C-terminus

详细信息    查看全文

• 作者：Jason C. Bermak ; Ming Li ; Clayton Bullock ; Paul Weingarten ; Qun-Yong Zhou
• 关键词：Biogenesis ; coatomer protein ; dopamine receptors ; mutagenesis ; endoplasmic reticulum
• 刊名：European Journal of Cell Biology
• 出版年：2002
• 出版时间：February 2002
• 年：2002
• 卷：81
• 期：2
• 页码：77-85
• 全文大小：309 K

文摘

Truncations at the carboxyl termini of G protein-coupled receptors result in defective receptor biogenesis and comprise a number of inherited disorders. In order to evaluate the structural role of the C-terminus in G protein-coupled receptor biogenesis, we generated a series of deletion and substitution mutations in the dopamine D1 receptor and visualized receptor subcellular localization by fusion to a green fluorescent protein. Alanine substitutions of several hydrophobic residues within the proximal C-terminus resulted in receptor transport arrest in the ER. Agonist binding and coupling to adenylyl cyclase was also abolished. In contrast, substitutions conserving C-terminal hydrophobicity produced normal cell surface receptor expression, binding, and stimulatory function. A mechanism for the role of the C-terminus in D1 receptor transport was investigated by searching for candidate protein interactions. The D1 receptor was found to co-precipitate and associate in vitro directly with the γ-subunit of the COPI coatomer complex. In vitro pull-down assays confirmed that only the D1 C-terminus is required for COPI association, and that identical mutations causing disruption of receptor transport to the cell surface also disrupted binding to COPI. Furthermore, conservative mutations in the D1 C-terminus restored COPI association just as they restored cell surface transport. These results suggest that association between the coatomer complex and hydrophobic residues within the proximal C-terminus of the D1 receptor may serve an important role in receptor transport.