Identification of G protein alpha subunit-palmitoylating enzyme.
The heterotrimeric G protein alpha subunit (Galpha) is targeted to the cytoplasmic face of the plasma membrane through reversible lipid palmitoylation and relays signals from G-protein-coupled receptors (GPCRs) to its effectors. By screening 23 DHHC motif (Asp-His-His-Cys) palmitoyl acyl-transferases, we identified DHHC3 and DHHC7 as Galpha palmitoylating enzymes. DHHC3 and DHHC7 robustly palmitoylated Galpha(q), Galpha(s), and Galpha(i2) in HEK293T cells. Knockdown of DHHC3 and DHHC7 decreased Galpha(q/11) palmitoylation and relocalized it from the plasma membrane into the cytoplasm. Photoconversion analysis revealed that Galpha(q) rapidly shuttles between the plasma membrane and the Golgi apparatus, where DHHC3 specifically localizes. Fluorescence recovery after photobleaching studies showed that DHHC3 and DHHC7 are necessary for this continuous Galpha(q) shuttling. Furthermore, DHHC3 and DHHC7 knockdown blocked the alpha(1A)-adrenergic receptor/Galpha(q/11)-mediated signaling pathway. Together, our findings revealed that DHHC3 and DHHC7 regulate GPCR-mediated signal transduction by controlling Galpha localization to the plasma membrane.
Tsutsumi
Ryouhei
R
Division of Membrane Physiology, Department of Cell Physiology, National Institute for Physiological Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan.
Fukata
Yuko
Y
Noritake
Jun
J
Iwanaga
Tsuyoshi
T
Perez
Franck
F
Fukata
Masaki
M
D000217:Acyltransferases / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D036681:Fluorescence Recovery After Photobleaching; D051960:GTP-Binding Protein alpha Subunit, Gi2 / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism*; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006056:Golgi Apparatus / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D054878:Lipoylation; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism; D021381:Protein Transport / Q000235:genetics; D034741:RNA, Small Interfering / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics
D034741:RNA, Small Interfering; D043562:Receptors, G-Protein-Coupled; D000217:Acyltransferases; D051960:GTP-Binding Protein alpha Subunit, Gi2; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D019205:GTP-Binding Protein alpha Subunits, Gs