Role of protein kinase D (PKD) phosphorylation of Oxysterol binding protein (OSBP) in sphingomyelin biosynthesis

Abstract

Oxysterol binding protein (OSBP) is a high affinity receptor for oxysterols that interacts with the endoplasmic reticulum (ER) and trans‐Golgi network via FFAT and pleckstrin homology (PH) domains, respectively. In vitro, OSBP displays cholesterol extraction and transfer activity and regulates the cholesterol content of Golgi and post‐Golgi membranes. Through this activity, OSBP controls CERT‐mediated ER‐to‐Golgi ceramide transfer, which is the rate‐determining step in sphingomyelin (SM) biosynthesis. CERT and OSBP are phosphorylated by protein kinase D (PKD), thus we investigated whether CERT activity and SM synthesis is regulated by OSBP phosphorylation at serine 242 by PKD. OSBP and PKD are physically associated, however OSBP is not required for Golgi association and activation of PKD. While transfection of wild‐type OSBP restored sterol activation of SM biosynthesis in OSBP‐depleted CHO cells, OSBP S242A and S242D were ineffective. Another OSBP‐related protein ORP9, which transfers cholesterol from ER to Golgi but is not involved in SM biosynthesis, is not a substrate for or associated with PKD. We conclude that an OSBP phosphorylation/dephosphorylation cycle mediated by PKD is necessary and specific for OSBP function in the context of regulating SM biosynthesis. Supported by the Canadian Institutes of Health Research.