Multiple forms of the U2 small nuclear ribonucleoprotein auxiliary factor U2AF subunits expressed in higher plants

Requirements for intron recognition during pre-mRNA splicing in plants diff er from those in vertebrates and yeast. Plant introns contain neither conse rved branch points nor distinct 3' splice site-proximal polypyrimidine trac ts characteristic of the yeast and vertebrate introns, respectively. Howeve r, they are strongly enriched in U residues throughout the intron, property essential for splicing. To understand the roles of different sequence elem ents in splicing, we are characterizing proteins involved in intron recogni tion in plants. In this work we show that Nicotiana plumbaginifolia, a dico tyledonous plant, contains two genes encoding different homologs of the lar ge 50-65-kDa subunit of the polypyrimidine tract binding factor U2AF, chara cterized previously in animals and Schizosaccharomyces pombe. Both plant U2 AF(65) isoforms, referred to as NpU2AF(65)a and NpU2AF(65)b, support splici ng of an adenovirus pre-mRNA in HeLa cell nuclear extracts depleted of the endogenous U2AF factor. Both proteins interact with RNA fragments containin g plant introns and show affinity for poly(U) and, to a lesser extend, poly (C) and poly(G). The branch point or the 3' splice site regions do not cont ribute significantly to intron recognition by NpU2AF(65). The existence of multiple isoforms of U2AF may be quite general in plants because two genes expressing U2AF(65) have been identified in Arabidopsis, and different isof orms of the U2AF small subunit are expressed in rice.