Diacylglycerol kinase delta suppresses ER-to-Golgi traffic via its SAM and PH domains

• Published in: Molecular biology of the cell 2002-01, Vol.13 (1), p.302-316
• Main Authors: Nagaya, Hisao, Wada, Ikuo, Jia, Yan-Jun, Kanoh, Hideo
• Format: Article
• Language: English
• Subjects: 3T3 Cells; Amino Acid Sequence; Animals; Blood Proteins - chemistry; Brefeldin A - pharmacology; Cell Line; COP-Coated Vesicles - metabolism; COS Cells; Diacylglycerol Kinase - chemistry; Diacylglycerol Kinase - genetics; Diacylglycerol Kinase - metabolism; Endoplasmic Reticulum - drug effects; Endoplasmic Reticulum - metabolism; Epithelial Cells - metabolism; Golgi Apparatus - drug effects; Golgi Apparatus - metabolism; Green Fluorescent Proteins; Intracellular Membranes - metabolism; Luminescent Proteins - analysis; Mice; Microscopy, Confocal; Microsomes, Liver - metabolism; Phosphoproteins - chemistry; Point Mutation; Protein Structure, Tertiary; Protein Transport - drug effects; Rats; Recombinant Fusion Proteins - metabolism; Vesicular stomatitis virus
• ISSN: 1059-1524; 1939-4586
• DOI: 10.1091/mbc.01-05-0255

We report here that the anterograde transport from the endoplasmic reticulum (ER) to the Golgi was markedly suppressed by diacylglycerol kinase delta (DGKdelta) that uniquely possesses a pleckstrin homology (PH) and a sterile alpha motif (SAM) domain. A low-level expression of DGKdelta in NIH3T3 cells caused redistribution into the ER of the marker proteins of the Golgi membranes and the vesicular-tubular clusters (VTCs). In this case DGKdelta delayed the ER-to-Golgi traffic of vesicular stomatitis virus glycoprotein (VSV G) and also the reassembly of the Golgi apparatus after brefeldin A (BFA) treatment and washout. DGKdelta was demonstrated to associate with the ER through its C-terminal SAM domain acting as an ER-targeting motif. Both of the SAM domain and the N-terminal PH domain of DGKdelta were needed to exert its effects on ER-to-Golgi traffic. Kinase-dead mutants of DGKdelta were also effective as the wild-type enzyme, suggesting that the catalytic activity of DGK was not involved in the present observation. Remarkably, the expression of DGKdelta abrogated formation of COPII-coated structures labeled with Sec13p without affecting COPI structures. These findings indicate that DGKdelta negatively regulates ER-to-Golgi traffic by selectively inhibiting the formation of ER export sites without significantly affecting retrograde transport.