Asymmetric dimethylation of AMPKa1 by PRMT6 contributes to the formation of phase-separated puncta

AMP-activated protein kinase (AMPK) is a heterotrimeric serine/threonine kinase comprising a, 13, and g subunits. AMPK is involved in intracellular energy metabolism and functions as a switch that turns various biological pathways in eukaryotes on and off. Several post-translational modifications regulating AMPK function have been demonstrated, including phosphorylation, acetylation, and ubiquitination; however, arginine methylation has not been reported in AMPKa1. We investigated whether arginine methylation occurs in AMPKa1. Screening experiments revealed arginine methylation of AMPKa1 mediated by protein arginine methyltransferase 6 (PRMT6). In vitro methylation and co-immunoprecipitation assays indicated that PRMT6 can directly interact with and methylate AMPKa1 without involvement of other intracellular components. In vitro methylation assays with truncated and point mutants of AMPKa1 revealed that Arg403 is the residue methylated by PRMT6. Immunocyto-chemical studies showed that the number of AMPKa1 puncta was enhanced in saponin-permeabilized cells when AMPKa1 was co-expressed with PRMT6, suggesting that PRMT6-mediated methylation of AMPKa1 at Arg403 alters the physiological characteristics of AMPKa1 and may lead to liquid-liquid phase separation.(c) 2023 Published by Elsevier Inc.