Identification and characterization of proteins that interact with the carboxy terminus of poly(A)-binding protein and inhibit translation in vitro.
Poly(A)-binding proteins (PABPs) are multifunctional proteins that play important roles in mRNA stability and protein translation. Two cucumber ( Cucumis sativus L.) proteins, PCI6 (PABP-CT-interacting) and PCI243 were identified based on ability to interact with the carboxy terminus (CT) of PABP in yeast two-hybrid and in vitro binding assays. PCI6 and PCI243 share a conserved amino acid domain (SxLnpnApxFxP) in common with human PABP-CT interactors, and with Arabidopsis ERD15 (early-responsive to dehydration). Deletion analysis and point mutations indicate that presence of this domain is necessary for the interaction, and tests with ERD15 demonstrate that it is predictive of interaction. Other plant proteins possessing this domain fall into two categories: small, acidic proteins like PCI6, PCI243 and ERD15, and larger neutral proteins that also include an RNA recognition motif. PCI6 is expressed in a range of tissues, e.g., leaves, roots, stems and flowers, and follows a diurnal pattern of expression, increasing during light hours and declining overnight. In wheat germ and mouse ascites Krebs-2 in vitro translation systems, PCI6 inhibited translation whereas the non-interacting mutant, PCI6-23A, did not or had a greatly reduced effect. The activity of PCI6, therefore, is reminiscent of that of human PABP-interacting protein 2 (Paip2). These results demonstrate a novel interaction between PABP and several plant proteins sharing a SxLnpxApxFxP motif, with possible implications for translational regulation.
Wang
Xiaofeng
X
Genetics Program, 342 Plant and Soil Sciences Building, Michigan State University, East Lansing, MI 48824, USA.
Grumet
Rebecca
R
D000595:Amino Acid Sequence; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D015152:Blotting, Northern; D003001:Cloning, Molecular; D018553:Cucumis sativus / Q000235:genetics / Q000378:metabolism; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D018506:Gene Expression Regulation, Plant; D008969:Molecular Sequence Data; D009154:Mutation; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D039102:Poly(A)-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D014176:Protein Biosynthesis / Q000235:genetics*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques
D029681:Arabidopsis Proteins; D018076:DNA, Complementary; C441601:ERD15 protein, Arabidopsis; D010940:Plant Proteins; D039102:Poly(A)-Binding Proteins