The assembly of the CAAT-box binding complex at a photosynthesis gene promoter is regulated by light, cytokinin, and the stage of the plastids

A functionally important region in the promoter of the spinach photosynthes is gene AtpC, which encodes the subunit gamma of the chloroplast ATP syntha se, is located immediately upstream of the CAAT-box, A single nucleotide ex change in this region (AAATTCAAT --> AAGATCAAT) uncouples the expression of an AtpC promoter::uidA gene fusion from the regulation by light, cytokinin , and functional plastids and results in a high constitutive expression of the reporter gene. By screening an Arabidopsis thaliana expression library with a double-stranded wild-type oligonucleotide from this promoter region, we have isolated cDNAs from Arabidopsis libraries that code for plant homo logs of the CAAT-box binding factor (CBF)-C, Binding occurs only in the pre sence of nuclear extracts, consistent with reports from metazoa CBFs that t he subunits A and B in addition to C are required for the formation of the CBF-DNA complex. At least eight genes with homologies to CBF-C are present in the Arabidopsis genome; one of them exhibits striking similarities to th e gene for the human global transcriptional repressor Drap1, In gel mobilit y shift assays, low binding activity of CBF to the wild-type AtpC promoter sequence was observed with nuclear extracts from tissue with low AtpC expre ssion levels, i.e. extracts from etiolated and photobleached seedlings, whe reas high binding activity was detectable with extracts from tissues with h igh AtpC expression levels, i.e. extracts from light-grown seedlings and et iolated seedlings treated with cytokinin, Binding to the mutant sequence, w hich directs constitutive high level uidA expression in vivo, is significan tly stronger than to the wild-type sequence. The data are consistent with t he idea that the assembly of CBF at the AtpC promoter is regulated in respo nse to light and cytokinin and that the low level of expression in etiolate d and photobleached material is caused by an inhibitory effect. The structu re/function relationships of the Arabidopsis CBFs are discussed in relation to their regulatory function in AtpC gene expression.