Caveolin-1 - A Novel Interacting Partner of Organic Cation/Carnitine Transporter (Octn2): Effect of Protein Kinase C on This Interaction in Rat Astrocytes

Abstract

OCTN2 - the Organic Cation Transporter Novel family member 2 (SLC22A5) is known to be a xenobiotic/drug transporter. It transports as well carnitine - a compound necessary for oxidation of fatty acids and mutations of its gene cause primary carnitine deficiency. Octn2 regulation by protein kinase C (PKC) was studied in rat astrocytes - cells in which β-oxidation takes place in the brain. Activation of PKC with phorbol ester stimulated L-carnitine transport and increased cell surface presence of the transporter, although no PKC-specific phosphorylation of Octn2 could be detected. PKC activation resulted in an augmented Octn2 presence in cholesterol/sphingolipid-rich microdomains of plasma membrane (rafts) and increased co-precipitation of Octn2 with raft-proteins, caveolin-1 and flotillin-1. Deletion of potential caveolin-1 binding motifs pointed to amino acids 14–22 and 447–454 as the caveolin-1 binding sites within Octn2 sequence. A direct interaction of Octn2 with caveolin-1 in astrocytes upon PKC activation was detected by proximity ligation assay, while such an interaction was excluded in case of flotillin-1. Functioning of a multi-protein complex regulated by PKC has been postulated in rOctn2 trafficking to the cell surface, a process which could be important both under physiological conditions, when carnitine facilitates fatty acids catabolism and controls free Coenzyme A pool as well as in pathology, when transport of several drugs can induce secondary carnitine deficiency.

Functional Expression of Organic Ion Transporters in Astrocytes and Their Potential as a Drug Target in the Treatment of Central Nervous System Diseases

“…This has been further supported by subsequent reports of OCTN2 in rats 40,41) and humans. 23) The presence of OCTN2 in astrocytes seems reasonable, since astrocytes require substantial amounts of energy to fulfill their roles.…”

Protein Kinases C-Mediated Regulations of Drug Transporter Activity, Localization and Expression

“…By contrast, PKCζ activation leads to increased OAT1 and OAT3 activity in rodent renal cortical slices, which may result, at least for OAT3, from increased trafficking into the plasma membrane [128]. With respect to the carnitine transporter OCTN2, its presence at the cell surface, as well as its activity, have been shown to be enhanced by PMA in cultured rat astrocytes, thus supporting the idea of a multi-protein complex regulated by PKCs and implicated in OCTN2 trafficking to the cell surface [72]. …”

l-Carnitine and Acetyl-l-carnitine Roles and Neuroprotection in Developing Brain

“…Uptake into brain also occurs primarily via the OCTN2 transporter [1, 44, 45]. There is evidence that the B 0,+ transporter, a Na + /Cl − dependent amino acid/carnitine transporter, also has a role in L-carnitine and acetyl-L-carnitine uptake into brain [44, 46–48], and into astrocytes [49, 50]. Immunohistochemical studies show that labeling for OCTN 1, 2 and 3 is distributed in many regions of mouse brain and spinal cord in a pattern consistent with possible roles in modulating bioenergetics and cholinergic neurotransmission [51].…”