RBP45 and RBP47, two oligouridylate-specific hnRNP-like proteins interacting with poly(A)+ RNA in nuclei of plant cells.
Introns in plant nuclear pre-mRNAs are highly enriched in U or U + A residues and this property is essential for efficient splicing. Moreover, 3'-untranslated regions (3'-UTRs) in plant pre-mRNAs are generally UA-rich and contain sequences that are important for the polyadenylation reaction. Here, we characterize two structurally related RNA-binding proteins (RBPs) from Nicotiana plumbaginifolia, referred to as RBP45 and RBP47, having specificity for oligouridylates. Both proteins contain three RBD-type RNA-binding domains and a glutamine-rich N-terminus, and share similarity with Nam8p, a protein associated with U1 snRNP in the yeast Saccharomyces cerevisiae. Deletion analysis of RBP45 and RBP47 indicated that the presence of at least two RBD are required for interaction with RNA and that domains other than RBD do not significantly contribute to binding. mRNAs for RBP45 and RBP47 and mRNAs encoding six related proteins in Arabidopsis thaliana are constitutively expressed in different plant organs. Indirect immunofluorescence and fractionation of cell extracts showed that RBP45 and RBP47 are localized in the nucleus. In vivo UV crosslinking experiments demonstrated their association with the nuclear poly(A)+ RNA. In contrast to UBP1, another oligouridylate-binding nuclear three-RBD protein of N. plumbaginifolia (Lambermon et al., EMBO J, 2000, 19:1638-1649), RBP45 and RBP47 do not stimulate mRNA splicing and accumulation when transiently overexpressed in protoplasts. Properties of RBP45 and RBP47 suggest they represent hnRNP-proteins participating in still undefined steps of pre-mRNA maturation in plant cell nuclei.
Lorković
Z J
ZJ
Friedrich Miescher-Institut, Basel, Switzerland.
Wieczorek Kirk
D A
DA
Klahre
U
U
Hemmings-Mieszczak
M
M
Filipowicz
W
W
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites; D002467:Cell Nucleus / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli; D019143:Evolution, Molecular; D008969:Molecular Sequence Data; D009843:Oligoribonucleotides / Q000378:metabolism*; D010802:Phylogeny; D010940:Plant Proteins; D010947:Plants, Toxic; D012333:RNA, Messenger / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014026:Nicotiana / Q000235:genetics* / Q000378:metabolism; D014500:Uracil Nucleotides / Q000378:metabolism*
D009843:Oligoribonucleotides; D010940:Plant Proteins; C417026:RBP45 protein, Nicotiana plumbaginifolia; C417027:RBP47 protein, Nicotiana plumbaginifolia; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; D014500:Uracil Nucleotides; C027916:oligo(U)