title: CmRBP50 protein phosphorylation is essential for assembly of a stable phloem-mobile high-affinity ribonucleoprotein complex.
abstract: RNA-binding proteins (RBPs) form ribonucleoprotein (RNP) complexes that play crucial roles in RNA processing for gene regulation. The angiosperm sieve tube system contains a unique population of transcripts, some of which function as long-distance signaling agents involved in regulating organ development. These phloem-mobile mRNAs are translocated as RNP complexes. One such complex is based on a phloem RBP named Cucurbita maxima RNA-binding protein 50 (CmRBP50), a member of the polypyrimidine track binding protein family. The core of this RNP complex contains six additional phloem proteins. Here, requirements for assembly of this CmRBP50 RNP complex are reported. Phosphorylation sites on CmRBP50 were mapped, and then coimmunoprecipitation and protein overlay studies established that the phosphoserine residues, located at the C terminus of CmRBP50, are critical for RNP complex assembly. In vitro pull-down experiments revealed that three phloem proteins, C. maxima phloem protein 16, C. maxima GTP-binding protein, and C. maxima phosphoinositide-specific phospholipase-like protein, bind directly with CmRBP50. This interaction required CmRBP50 phosphorylation. Gel mobility-shift assays demonstrated that assembly of the CmRBP50-based protein complex results in a system having enhanced binding affinity for phloem-mobile mRNAs carrying polypyrimidine track binding motifs. This property would be essential for effective long-distance translocation of bound mRNA to the target tissues.
authors.affiliation: Department of Plant Biology, College of Biological Sciences, University of California, Davis, California, 95616, USA.
authors.forename: Pingfang
authors.initials: P
authors.lastname: Li
authors.forename: Byung-Kook
authors.initials: BK
authors.lastname: Ham
authors.forename: William J
authors.initials: WJ
authors.lastname: Lucas
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D028464:Cucurbita / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D052585:Phloem / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000502:physiology; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D018749:RNA, Plant / Q000235:genetics / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000235:genetics / Q000378:metabolism*; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism
chemical_list: D010940:Plant Proteins; D012333:RNA, Messenger; D018749:RNA, Plant; D012261:Ribonucleoproteins