Diacylglycerol kinase (DGK) is a lipid kinase to convert DG to phosphatidic acid. Recently, we have reported that a-tocopherol (αToc) and epigallocatechin gallate (EGCG) activate DGKα via 67kDa laminin receptor (67LR), resulting in improvement of diabetic nephropathy. Interestingly, αToc directly binds to 67LR at different site for EGCG, suggesting unknown binding site of αToc in 67LR. Furthermore, 67LR is believed to be concentrated in lipid raft as homo dimer. However, palmitoylation of 67LR hasn’t been reported, although many proteins localized in raft are palmitoylated. Therefore, we investigated αToc binding site of 67LR and palmitoylation of 67LR with its physiological meaning in the activation of DGKα.
We identified the αToc binding site in 67LR using hydrogen-deuterium exchange mass spectrometry, and found both EGCG and αToc treatment induced palmitoylation with DGKα translocation (activation). A time course of αToc-induced palmitoylation was similar with DGKα translocation. A point mutation at Cys of 67LR abolished both 67LR palmitoylation and DGKα activation. These results indicate that palmitoylation of 67LR is necessary for the activation of DGKα by αToc or EGCG, suggested that, as well as DGKα, 67LR and its palmitoylating enzyme are pharmaceutical targets of diabetic nephropathy.