Redox and heavy metal effects on the biochemical activities of an Arabidopsis polyadenylation factor subunit.
The Arabidopsis CPSF30 ortholog (AtCPSF30) is an RNA-binding endonuclease that is part of the plant polyadenylation complex. Previous work (B. Addepalli, A.G. Hunt, Nucleic Acids Res. 35 (2007) 4453-4463) demonstrated that different zinc finger motifs in the protein were responsible for RNA-binding and nuclease activity, respectively. In this study, a more detailed functional map of AtCPSF30 is presented, a map that includes descriptions of novel biochemical activities. Elevated temperatures, the specific zinc chelator 1,10-phenanthroline, and the sulfhydryl reagent dithiothreitol all had differential inhibitory effects on the RNA-binding and nuclease activities. The endonuclease activity of AtCPSF30 was inhibited by relatively high (>100muM) concentrations of zinc, and this inhibition required a plant-specific N-terminal domain apart from the zinc finger core of the protein. ATP stimulated the nuclease activity in the presence of zinc, and this stimulation required a plant-specific C-terminal domain, again apart from the zinc finger core. These studies reveal a subtle and unexpected complexity to AtCPSF30, and raise the possibility that multiple avenues of regulation may impinge on this protein through different functional domains.
Addepalli
Balasubrahmanyam
B
Department of Plant and Soil Sciences, University of Kentucky, 301A Plant Science Building, 1405 Veterans Drive, Lexington, KY 40546-0312, USA.
Hunt
Arthur G
AG
D000255:Adenosine Triphosphate / Q000737:chemistry; D000595:Amino Acid Sequence; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002413:Cations, Divalent / Q000737:chemistry; D039223:Cleavage And Polyadenylation Specificity Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004252:DNA Mutational Analysis; D004229:Dithiothreitol / Q000737:chemistry; D019216:Metals, Heavy / Q000737:chemistry*; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D012492:Salts / Q000737:chemistry; D013439:Sulfhydryl Reagents / Q000737:chemistry; D013696:Temperature; D016335:Zinc Fingers
D029681:Arabidopsis Proteins; C521936:CPSF30 protein, Arabidopsis; D002413:Cations, Divalent; D039223:Cleavage And Polyadenylation Specificity Factor; D019216:Metals, Heavy; D021122:Protein Subunits; D012492:Salts; D013439:Sulfhydryl Reagents; D000255:Adenosine Triphosphate; D004229:Dithiothreitol