Diacylglycerol kinase delta suppresses ER-to-Golgi traffic via its SAM and PH domains.
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.
Nagaya
Hisao
H
Department of Biochemistry, Sapporo Medical University School of Medicine, Sapporo 060-8556, Japan.
Wada
Ikuo
I
Jia
Yan-Jun
YJ
Kanoh
Hideo
H
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001798:Blood Proteins / Q000737:chemistry; D020126:Brefeldin A / Q000494:pharmacology; D022181:COP-Coated Vesicles / Q000378:metabolism; D019556:COS Cells; D002460:Cell Line; D019852:Diacylglycerol Kinase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D004847:Epithelial Cells / Q000378:metabolism; D006056:Golgi Apparatus / Q000187:drug effects / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D007425:Intracellular Membranes / Q000378:metabolism; D008164:Luminescent Proteins / Q000032:analysis; D051379:Mice; D018613:Microscopy, Confocal; D008862:Microsomes, Liver / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000187:drug effects; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D001798:Blood Proteins; D008164:Luminescent Proteins; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; C039077:platelet protein P47; D049452:Green Fluorescent Proteins; D020126:Brefeldin A; D019852:Diacylglycerol Kinase