Phosphorylation of nuclear proteins directs binding to salicylic acid-responsive elements.
The cis-located DNA sequence as-1 (Activation Sequence-1) from CaMV 35S promoter has been previously identified as an element that can confer inducibility by salicylic acid (SA) with immediate early kinetics. This sequence specifically binds to ASF-1 (Activation Sequence Factor-1), previously characterized in tobacco nuclear extracts. To assess whether modulation of ASF-1 binding activity can explain the activation of the as-1 sequence observed in vivo, we performed electrophoretic mobility shift assays using nuclear extracts from SA-treated and water-treated tobacco plants. Our results indicate that treatment of plants with SA increases ASF-1 binding to as-1 and to ocs, an as-1-like element from the Agrobacterium octopine synthase gene. In contrast, SA treatment has no effect on the binding of GT-1 factor to its target light-inducible box II element. Furthermore, treatment of nuclear extracts from SA-treated plants with alkaline phosphatase decreases ASF-1 binding to the as-1 element. This can be reversed by pretreatment with 10 mM NaF. Accordingly, pretreatment of nuclear extracts from control water-treated plants with ATP produces an increase in ASF-1 binding activity similar to that observed with SA. This effect of ATP is reversed by treatment with alkaline phosphatase and prevented by quercetin, a casein kinase II inhibitor. These results support the hypothesis that a nuclear protein kinase is involved in the immediate early events of transcriptional activation triggered by SA.
Stange
C
C
Departamento de Genética Molecular y Microbiología, Facultad de Ciencias Biológicas, Pontificia Universidad Católica de Chile, Santiago, Chile.
Ramírez
I
I
Gómez
I
I
Jordana
X
X
Holuigue
L
L
D029681:Arabidopsis Proteins; D050976:Basic-Leucine Zipper Transcription Factors; D001665:Binding Sites; D018744:DNA, Plant / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D010766:Phosphorylation; D010940:Plant Proteins / Q000378:metabolism; D010947:Plants, Toxic; D012459:Salicylates / Q000378:metabolism*; D020156:Salicylic Acid; D014026:Nicotiana / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation
C084054:ASF-2 protein, tobacco; D029681:Arabidopsis Proteins; D050976:Basic-Leucine Zipper Transcription Factors; D018744:DNA, Plant; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D010940:Plant Proteins; D012459:Salicylates; C087770:TGA3 protein, Arabidopsis; D014157:Transcription Factors; D020156:Salicylic Acid