De Novo Activation of the β-Phaseolin Promoter by Phosphatase or Protein Synthesis Inhibitors

Abstract

The promoter for the phaseolin (phas) bean seed protein gene adopts an inactive chromatin structure in leaves of transgenic tobacco. This repressive architecture, which confers stringent spatial regulation, is disrupted upon transcriptional activation during embryogenesis in a process that requires the presence of both a transcription factor (PvALF) and abscisic acid (ABA). Toward determining the need for de novo synthesis of proteins other than PvALF in transcriptional activation we explored the effect of several eukaryotic protein synthesis inhibitors. Surprisingly, cycloheximide (CHX), emetine, and verrucarin A were able to induce transcription from the phas promoter in tobacco and bean leaf tissue in the absence of either PvALF or ABA. This induction was decreased by the replication inhibitors hydroxyurea and aphidicolin but not by genistein or mimosine. Since protein phosphatases and kinases are essential components of the ABA signal transduction pathway, it is conceivable that CHX is also capable of inducing phosphorylation of proteins usually involved in ABA-mediated activation. Interestingly, okadaic acid, an inhibitor of serine/threonine phosphatase, also strongly activated transcription from the phas promoter. In contrast, the protein synthesis inhibitors anisomycin and puromycin did not activate transcription from the phas promoter, nor did the tyrosine phosphatase inhibitors phenylarsine oxide and sodium orthovanadate. These discrete but different results on transcriptional activation may reflect specific modes of action of the inhibitors, or they may reflect differential interactions of the inhibitors or of downstream events resulting from inhibitor activity with presently unknown components of the transcriptional activation system.

Interaction of PvALF and VP1 B3 domains with the β-phaseolin promoter

“…These results, together with the modulespecific regulation of the phas promoter observed during embryogenesis (Chandrasekharan et al, 2003a) and likely intervention of a protein phosphatase (Li et al, 2001a), provide evidence that multiple factors participate coordinately in PvALF and ABA-mediated phas promoter activation. Since chromatin architecture is among these components, the possible effect of histones on binding of the B3 domain proteins to the phas promoter was investigated.…”

Nectar chemistry

“…Zauralov (1969) pounds secreted into nectar (see "Phenolics" on page 247). Transglucosidase phate starvation (Bozzo et al, 2002), signal transduction (Li et al, 2001),…”

Exogenously induced expression of ethylene biosynthesis, ethylene perception, phospholipase D, and Rboh-oxidase genes in broccoli seedlings

“…CHX has mostly been considered as a eukaryotic protein synthesis inhibitor, but this seems to be only one aspect of its action in living cells. According to some suggestions (Li et al , 2001), CHX can (i) lead to transcriptional activation via loss of labile negative regulators; (ii) inhibit translation of protein products of autorepressive genes, and thus superinduce their transcription; (iii) prevent the synthesis of labile mRNA-degrading enzymes; (iv) cause RNAs to be trapped on polysomes, thus shielding them from cytoplasmic RNases; (v) induce phosphorylation of proteins usually involved in abscisic acid (ABA)-mediated activation; (vi) lead to direct transcriptional activation via phosphorylation of H3 histones; and (vii) uncouple DNA replication and chromatin assembly preventing the formation of a repressive chromatin structure.…”