Molecular cloning and characterization of rho, a ras-related small GTP-binding protein from the garden pea.
The rho proteins, members of the ras superfamily of small GTP-binding proteins, play a central role in the modulation of cellular functions involving the actin cytoskeleton such as in the establishment of cell polarity and morphology. As a first step in elucidating signal transduction pathways leading to processes mediated by the actin cytoskeleton in plants, we initiated cloning and characterization of rho proteins from pea. One rho-related, partial cDNA clone of 167 bp was isolated utilizing a polymerase chain reaction-based cloning strategy, using degenerate primers that correspond to conserved domains within the rho proteins. A full-length cDNA was isolated by screening a pea cDNA library using the 167-bp cDNA as a probe. The Rho1Ps cDNA contains an open reading frame encoding a polypeptide (Rho1Ps) of 197 amino acids that shows 45-64% sequence identity to members of the rho family and about 30% identity to other members of the ras superfamily. In addition to the nucleotide-binding and GTPase domains, Rho1Ps shares conserved residues and motifs unique to the rho proteins. Purified Rho1Ps protein expressed in Escherichia coli retains specific GTP-binding activity. These data indicate that Rho1Ps encodes a small GTP-binding protein of the rho family. The Rho1Ps transcript is expressed in all organs of pea seedlings, being more abundant in root tips and apical buds. DNA gel blot analyses show that the rho proteins in pea are encoded by a multigene family.
Yang
Z
Z
Department of Botany, University of Maryland, College Park 20742-5815.
Watson
J C
JC
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D007887:Fabaceae / Q000235:genetics / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D017343:Genes, Plant; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D005810:Multigene Family; D009838:Oligodeoxyribonucleotides; D010802:Phylogeny; D010940:Plant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D010946:Plants, Medicinal; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid; D020741:rho GTP-Binding Proteins
D009838:Oligodeoxyribonucleotides; D010940:Plant Proteins; C083285:RHO1 protein, Pisum sativum; D011993:Recombinant Fusion Proteins; D006160:Guanosine Triphosphate; D004247:DNA; D019204:GTP-Binding Proteins; D020741:rho GTP-Binding Proteins