Association of Arabidopsis type-II ROPs with the plasma membrane requires a conserved C-terminal sequence motif and a proximal polybasic domain.
Plant ROPs (or RACs) are soluble Ras-related small GTPases that are attached to cell membranes by virtue of the post-translational lipid modifications of prenylation and S-acylation. ROPs (RACs) are subdivided into two major subgroups called type-I and type-II. Whereas type-I ROPs terminate with a conserved CaaL box and undergo prenylation, type-II ROPs undergo S-acylation on two or three C-terminal cysteines. In the present work we determined the sequence requirement for association of Arabidopsis type-II ROPs with the plasma membrane. We identified a conserved sequence motif, designated the GC-CG box, in which the modified cysteines are flanked by glycines. The GC-CG box cysteines are separated by five to six mostly non-polar residues. Deletion of this sequence or the introduction of mutations that change its nature disrupted the association of ROPs with the membrane. Mutations that changed the GC-CG box glycines to alanines also interfered with membrane association. Deletion of a polybasic domain proximal to the GC-CG box disrupted the plasma membrane association of AtROP10. A green fluorescent protein fusion protein containing the C-terminal 25 residues of AtROP10, including its polybasic domain and GC-CG box, was primarily associated with the plasma membrane but a similar fusion protein lacking the polybasic domain was exclusively localized in the soluble fraction. These data provide evidence for the minimal sequence required for plasma membrane association of type-II ROPs in Arabidopsis and other plant species.
Lavy
Meirav
M
Department of Plant Sciences, Tel Aviv University, Tel Aviv 69978, Israel.
Yalovsky
Shaul
S
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D017124:Conserved Sequence; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D008969:Molecular Sequence Data; D020559:Monomeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary
D029681:Arabidopsis Proteins; D019204:GTP-Binding Proteins; C472133:ROP10 protein, Arabidopsis; D020559:Monomeric GTP-Binding Proteins; C512796:ROP9 protein, Arabidopsis