The Assembly of the CAAT-box Binding Complex at a Photosynthesis Gene Promoter Is Regulated by Light, Cytokinin, and the Stage of the Plastids

Abstract

A functionally important region in the promoter of the spinach photosynthesis gene AtpC, which encodes the subunit ␥ of the chloroplast ATP synthase, is located immediately upstream of the CAAT-box. A single nucleotide exchange in this region (AAAATTCAAT 3 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 homologs of the CAAT-box binding factor (CBF)-C. Binding occurs only in the presence of nuclear extracts, consistent with reports from metazoa CBFs that the 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 the gene for the human global transcriptional repressor Drap1. In gel mobility shift assays, low binding activity of CBF to the wild-type AtpC promoter sequence was observed with nuclear extracts from tissue with low AtpC expression levels, i.e. extracts from etiolated and photobleached seedlings, whereas high binding activity was detectable with extracts from tissues with high AtpC expression levels, i.e. extracts from light-grown seedlings and etiolated seedlings treated with cytokinin. Binding to the mutant sequence, which directs constitutive high level uidA expression in vivo, is significantly stronger than to the wild-type sequence. The data are consistent with the idea that the assembly of CBF at the AtpC promoter is regulated in response to light and cytokinin and that the low level of expression in etiolated and photobleached material is caused by an inhibitory effect. The structure/ function relationships of the Arabidopsis CBFs are discussed in relation to their regulatory function in AtpC gene expression.

Nuclear factor Y-mediated H3K27me3 demethylation of the SOC1 locus orchestrates flowering responses of Arabidopsis

“…Several studies have tested in vitro binding sites of plant NF-Y subunits 55,58 , but whether these sites are relevant to NF-Y endogenous function is unclear. Another recent study has proposed that NF-Y complexes may recognize a distal CCAAT site at the FT promoter based on ChIP assays of 35S:NF-YB2-YFP/HA plants 59 , whereas we were unable to detect binding of NF-YB2-6Myc to FT using the nf-yb2-1 pNF-YB2:NF-YB2-6Myc tagging line (Supplementary Fig.…”

The Promiscuous Life of Plant NUCLEAR FACTOR Y Transcription Factors

“…The initial reports of NF-Y genes in plants date back to the 1990s (Li et al, 1992b;Albani and Robert, 1995;Edwards et al, 1998;Lotan et al, 1998;Kusnetsov et al, 1999). The comprehensive search for NF-Y genes in Arabidopsis thaliana (Gusmaroli et al, 2001(Gusmaroli et al, , 2002Yang et al, 2005;Siefers et al, 2009) was followed by similar ones in other plant species, including rice (Oryza sativa), wheat (Triticum aestivum), and Brachypodium distachyon (Stephenson et al, 2007;Thirumurugan et al, 2008;Cao et al, 2011b).…”

LEAFY COTYLEDON1-LIKE Defines a Class of Regulators Essential for Embryo Development

“…Second, an AHAP2 gene has been shown to complement a yeast HAP2 mutation, indicating that the Arabidopsis protein can function in a CBF (Edwards et al, 1998). Third, an AHAP5 subunit has been shown to interact with other nuclear proteins, presumably AHAP2 and AHAP3, to form a complex that binds a double-stranded oligonucleotide containing a CAAT box (Kusnetsov et al, 1999). Fourth, loss-offunction mutations of L1L and LEC1, two HAP3 paralogs, have severe consequences on plant development, suggesting that these subunits play essential roles.…”

“…Although a CBF from plants has not been isolated, several lines of evidence suggest that L1L and LEC1 function as part of a CBF that regulates embryogenesis. First, in addition to HAP3, paralogs of the other two CBF subunits required for DNA binding activity, HAP2 and HAP5, have been identified in plants (Li et al, 1992;Albani and Robert, 1995;Edwards et al, 1998;Kusnetsov et al, 1999;Gusmaroli et al, 2001). Unlike yeast and mammals, which possess single genes for each subunit, plants possess families of subunits.…”