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Protein kinase D1 and oxysterol‐binding protein form a regulatory complex independent of phosphorylation
Author(s) -
Goto Asako,
Charman Mark,
Ridgway Neale D.
Publication year - 2018
Publication title -
traffic
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.677
H-Index - 130
eISSN - 1600-0854
pISSN - 1398-9219
DOI - 10.1111/tra.12609
Subject(s) - biology , phosphorylation , microbiology and biotechnology , oxysterol , protein kinase a , autophagy related protein 13 , protein phosphorylation , kinase , plasma protein binding , biochemistry , cholesterol
Protein kinase D (PKD) controls secretion from the trans‐Golgi network (TGN) by phosphorylating phosphatidylinositol 4‐kinase IIIβ and proteins that bind and/or transfer phosphatidylinositol 4‐phosphate (PtdIns‐4P), such as oxysterol‐binding protein (OSBP) and ceramide transfer protein. Here, we investigated the consequences of PKD phosphorylation of OSBP at endoplasmic reticulum (ER)‐Golgi membrane contact sites (MCS). Results with OSBP phospho‐mutants revealed that PKD phosphorylation did not affect sterol and PtdIns‐4P binding, activation of sphingomyelin (SM) synthesis at Golgi‐ER MCS or other OSBP phospho‐sites. Instead, an interaction was identified between the N‐terminal region of OSBP and PKD1 that was independent of kinase activity and OSBP phosphorylation status. S916 autophosphorylation of PKD1 was inhibited by OSBP expression suggesting the interaction negatively regulates PKD1 activity. Stimulation of PKD1 activity by phorbol ester promoted the Golgi‐localization of wild‐type and phospho‐mutants of OSBP but did not affect OSBP‐dependent SM synthesis. Only when wild‐type or kinase‐dead PKD1 was overexpressed was 25‐hydroxycholesterol‐activated SM synthesis inhibited. We conclude that OSBP and PKD1 form a complex that inhibits both the oxysterol‐dependent activity of OSBP at the ER‐Golgi and activation of PKD1. Formation of the complex was independent of PKD1 activity and phosphorylation of OSBP.