title: Structural basis for the Golgi association by the pleckstrin homology domain of the ceramide trafficking protein (CERT).
abstract: Ceramide transport from the endoplasmic reticulum to the Golgi apparatus is crucial in sphingolipid biosynthesis, and the process relies on the ceramide trafficking protein (CERT), which contains pleckstrin homology (PH) and StAR-related lipid transfer domains. The CERT PH domain specifically recognizes phosphatidylinositol 4-monophosphate (PtdIns(4)P), a characteristic phosphoinositide in the Golgi membrane, and is indispensable for the endoplasmic reticulum-to-Golgi transport of ceramide by CERT. In this study, we determined the three-dimensional structure of the CERT PH domain by using solution NMR techniques. The structure revealed the presence of a characteristic basic groove near the canonical PtdIns(4)P recognition site. An extensive interaction study using NMR and other biophysical techniques revealed that the basic groove coordinates the CERT PH domain for efficient PtdIns(4)P recognition and localization in the Golgi apparatus. The notion was also supported by Golgi mislocalization of the CERT mutants in living cells. The distinctive binding modes reflect the functions of PH domains, as the basic groove is conserved only in the PH domains involved with the PtdIns(4)P-dependent lipid transport activity but not in those with the signal transduction activity.
authors.affiliation: Graduate School of Pharmaceutical Sciences, University of Tokyo, Bunkyo-ku, Tokyo, Japan.
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Sugiki
authors.forename: Koh
authors.initials: K
authors.lastname: Takeuchi
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Yamaji
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Takano
authors.forename: Yuji
authors.initials: Y
authors.lastname: Tokunaga
authors.forename: Keigo
authors.initials: K
authors.lastname: Kumagai
authors.forename: Kentaro
authors.initials: K
authors.lastname: Hanada
authors.forename: Hideo
authors.initials: H
authors.lastname: Takahashi
authors.forename: Ichio
authors.initials: I
authors.lastname: Shimada
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008055:Lipids / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008958:Models, Molecular; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010716:Phosphatidylinositols / Q000737:chemistry; D011487:Protein Conformation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D020349:Surface Plasmon Resonance
chemical_list: D008055:Lipids; D008081:Liposomes; D010716:Phosphatidylinositols; C119090:CERT1 protein, human; D017346:Protein Serine-Threonine Kinases