Large scale interaction analysis predicts that the <Emphasis Type="Italic">Gerbera hybrida</Emphasis> floral E function is provided both by general and specialized proteins

Background  

The ornamental plant Gerbera hybrida bears complex inflorescences with morphologically distinct floral morphs that are specific to the sunflower family Asteraceae. We have previously characterized several MADS box genes that regulate floral development in Gerbera. To study further their behavior in higher order complex formation according to the quartet model, we performed yeast two- and three-hybrid analysis with fourteen Gerbera MADS domain proteins to analyze their protein-protein interaction potential.     

Minimal regions in the Arabidopsis PISTILLATA promoter responsive to the APETALA3/PISTILLATA feedback control do not contain a CArG box

PISTILLATA (PI) is a floral homeotic B function gene in Arabidopsis and together with the other B function gene, APETALA3 (AP3), is involved in specifying petal and stamen identities. The expression of PI and AP3 is under similar developmental control. The initiation of AP3 and PI expression is at least partly caused by the floral meristem identity gene LEAFY, but the maintenance of AP3 and PI expression involves an autoregulatory loop requiring the activity of both genes. PI and AP3 are MADS domain proteins that form, and appear to function as, a heterodimer. AP3/PI binds in vitro to a sequence motif, CC(A/T)₆GG, a MADS domain protein consensus binding site also known as the CArG box. We identified a 481-bp PI promoter region that confers both the initiation and the maintenance of PI expression patterns. We further dissected the promoter and identified minimal regions responsible for the AP3/PI-dependent expression. No CArG box is present in these minimal regions, suggesting that either AP3/PI does not bind directly to the PI promoter for the maintenance control, or that it requires additional factors to bind to the PI promoter. Our results suggest that the mechanisms of regulation of the two B function genes, AP3 and PI, are different, because CArG boxes are present in the AP3 promoter and are necessary for the AP3 feedback control. Received: 1 March 2000 / Revision accepted: 15 June 2000     

The Arabidopsis FLC protein interacts directly in vivo with SOC1 and FT chromatin and is part of a high-molecular-weight protein complex

The Arabidopsis Flowering Locus C (FLC) protein is a repressor of flowering regulated by genes in the autonomous and vernalization pathways. Previous genetic and transgenic data have suggested that FLC acts by repressing expression of the floral integrator genes SOC1 and FT. We have taken an in vivo approach to determine whether the FLC protein interacts directly with potential DNA targets. Using chromatin immunoprecipitation, we have shown that FLC binds to a region of the first intron of FT that contains a putative CArG box, and have confirmed that FLC binds to a CArG box in the promoter of the SOC1 gene. MADS box proteins are thought to bind their DNA targets as dimers or higher-order multimers. We have shown that FLC is a component of a multimeric protein complex in vivo and that more than one FLC polypeptides can be present in the complex.     

Determination of floral organ identity by Arabidopsis MADS domain homeotic proteins AP1, AP3, PI, and AG is independent of their DNA-binding specificity.

The MADS domain homeotic proteins APETALA1 (AP1), APETALA3 (AP3), PISTILLATA (PI), and AGAMOUS (AG) combinatorially specify the identity of Arabidopsis floral organs. AP1/AP1, AG/AG, and AP3/PI dimers bind to similar CArG box sequences; thus, differences in DNA-binding specificity among these proteins do not seem to be the origin of their distinct organ identity properties. To assess the overall contribution that specific DNA binding could make to their biological specificity, we have generated chimeric genes in which the amino-terminal half of the MADS domain of AP1, AP3, PI, and AG was substituted by the corresponding sequences of human SRF and MEF2A proteins. In vitro DNA-binding assays reveal that the chimeric proteins acquired the respective, and distinct, DNA-binding specificity of SRF or MEF2A. However, ectopic expression of the chimeric genes reproduces the dominant gain-of-function phenotypes exhibited by plants ectopically expressing the corresponding Arabidopsis wild-type genes. In addition, both the SRF and MEF2 chimeric genes can complement the pertinent ap1-1, ap3-3, pi-1, or ag-3 mutations to a degree similar to that of AP1, AP3, PI, and AG when expressed under the control of the same promoter. These results indicate that determination of floral organ identity by the MADS domain homeotic proteins AP1, AP3, PI, and AG is independent of their DNA-binding specificity. In addition, the DNA-binding experiments show that either one of the two MADS domains of a dimer can be sufficient to confer a particular DNA-binding specificity to the complex and that sequences outside the amino-terminal basic region of the MADS domain can, in some cases, contribute to the DNA-binding specificity of the proteins.