title: Individual palmitoyl residues serve distinct roles in H-ras trafficking, microlocalization, and signaling.
abstract: H-ras is anchored to the plasma membrane by two palmitoylated cysteine residues, Cys181 and Cys184, operating in concert with a C-terminal S-farnesyl cysteine carboxymethylester. Here we demonstrate that the two palmitates serve distinct biological roles. Monopalmitoylation of Cys181 is required and sufficient for efficient trafficking of H-ras to the plasma membrane, whereas monopalmitoylation of Cys184 does not permit efficient trafficking beyond the Golgi apparatus. However, once at the plasma membrane, monopalmitoylation of Cys184 supports correct GTP-regulated lateral segregation of H-ras between cholesterol-dependent and cholesterol-independent microdomains. In contrast, monopalmitoylation of Cys181 dramatically reverses H-ras lateral segregation, driving GTP-loaded H-ras into cholesterol-dependent microdomains. Intriguingly, the Cys181 monopalmitoylated H-ras anchor emulates the GTP-regulated microdomain interactions of N-ras. These results identify N-ras as the Ras isoform that normally signals from lipid rafts but also reveal that spacing between palmitate and prenyl groups influences anchor interactions with the lipid bilayer. This concept is further supported by the different plasma membrane affinities of the monopalmitoylated anchors: Cys181-palmitate is equivalent to the dually palmitoylated wild-type anchor, whereas Cys184-palmitate is weaker. Thus, membrane affinity of a palmitoylated anchor is a function both of the hydrophobicity of the lipid moieties and their spatial organization. Finally we show that the plasma membrane affinity of monopalmitoylated anchors is absolutely dependent on cholesterol, identifying a new role for cholesterol in promoting interactions with the raft and nonraft plasma membrane.
authors: Institute for Molecular Bioscience, 306 Carmody Road, University of Queensland, Brisbane 4072, Australia.
authors: Sandrine
authors: S
authors: Roy
authors: Sarah
authors: S
authors: Plowman
authors: Barak
authors: B
authors: Rotblat
authors: Ian A
authors: IA
authors: Prior
authors: Cornelia
authors: C
authors: Muncke
authors: Sarah
authors: S
authors: Grainger
authors: Robert G
authors: RG
authors: Parton
authors: Yoav I
authors: YI
authors: Henis
authors: Yoel
authors: Y
authors: Kloog
authors: John F
authors: JF
authors: Hancock
mesh_terms: D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D003545:Cysteine / Q000235:genetics; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D011905:Genes, ras; D006056:Golgi Apparatus / Q000378:metabolism; D007700:Kinetics; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism; D009154:Mutation; D015689:Oncogene Protein p21(ras) / Q000235:genetics / Q000378:metabolism*; D016716:PC12 Cells; D010169:Palmitic Acids / Q000378:metabolism*; D021381:Protein Transport / Q000235:genetics; D051381:Rats; D012694:Serine / Q000235:genetics; D048490:raf Kinases / Q000378:metabolism
chemical_list: D010169:Palmitic Acids; D012694:Serine; D048490:raf Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D020930:MAP Kinase Kinase Kinases; D015689:Oncogene Protein p21(ras); D003545:Cysteine