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 residu es and this property is essential for efficient splicing, Moreover, 3'-untr anslated regions (3'-UTRs) in plant pre-mRNAs are generally UA-rich and con tain sequences that are important for the polyadenylation reaction. Here, w e characterize two structurally related RNA-binding proteins (REPs) from Ni cotiana plumbaginifolia, referred to as RBP45 and RBP47, having specificity for oligouridylates. Both proteins contain three RED-type RNA-binding doma ins and a glutamine-rich N-terminus, and share similarity with Nam8p, a pro tein associated with U1 snRNP in the yeast Saccharomyces cerevisiae, Deleti on analysis of RBP45 and REP47 indicated that the presence of at least two RED are required for interaction with RNA and that domains other than RED d o not significantly contribute to binding, mRNAs for RBP45 and REP47 and mR NAs encoding six related proteins in Arabidopsis thaliana are constitutivel y expressed in different plant organs. Indirect immunofluorescence and frac tionation of cell extracts showed that REP45 and RBP47 are localized in the nucleus. In vivo UV crosslinking experiments demonstrated their associatio n with the nuclear poly(A)(+) RNA. In contrast to UBP1, another oligouridyl ate-binding nuclear three-RED protein of N, plumbaginifolia (Lambermon et a l., EMBO J, 2000, 19:1638-1649), RBP45 and RBP47 do not stimulate mRNA spli cing and accumulation when transiently overexpressed in protoplasts. Proper ties of RBP45 and RBP47 suggest they represent hnRNP-proteins participating in still undefined steps of pre-mRNA maturation in plant cell nuclei.