Discrete spatial and temporal cis-acting elements regulate transcription of the Arabidopsis floral homeotic gene APETALA3

Abstract

The APETALA3 floral homeotic gene is required for petal and stamen development in Arabidopsis. APETALA3 transcripts are first detected in a meristematic region that will give rise to the petal and stamen primordia, and expression is maintained in this region during subsequent development of these organs. To dissect how the APETALA3 gene is expressed in this spatially and temporally restricted domain, various APETALA3 promoter fragments were fused to the uidA reporter gene encoding beta-glucuronidase and assayed for the resulting patterns of expression in transgenic Arabidopsis plants. Based on these promoter analyses, we defined cis-acting elements required for distinct phases of APETALA3 expression, as well as for petal-specific and stamen-specific expression. By crossing the petal-specific construct into different mutant backgrounds, we have shown that several floral genes, including APETALA3, PISTILLATA, UNUSUAL FLORAL ORGANS, and APETALA1, encode trans-acting factors required for second-whorl-specific APETALA3 expression. We have also shown that the products of the APETALA1, APETALA3, PISTILLATA and AGAMOUS genes bind to several conserved sequence motifs within the APETALA3 promoter. We present a model whereby spatially and temporally restricted APETALA3 transcription is controlled via interactions between proteins binding to different domains of the APETALA3 promoter.

The F-box cofactor UFO redirects the LEAFY floral regulator to novel cis-elements

“…The molecular mechanism we discovered here is consistent with most published data on AP3 and PI regulation (Chae et al, 2008; Hill et al, 1998; Honma and Goto, 2000; Tilly et al, 1998). However, precise understanding of regulation of AP3 and RBE will require further work on other cis and trans -elements.…”

“…To decipher the molecular mechanism underlying the synergistic action of LFY and UFO, we used a Dual Luciferase Reporter Assay (DLRA) to analyze their ability to activate promoters when transiently expressed in Arabidopsis protoplasts. We found that pAP3, the best characterized LFY-UFO target (Hill et al, 1998; Lamb et al, 2002), is strongly activated when LFY (or LFY-VP16, a fusion of LFY with the VP16 activation domain; Parcy et al, 1998) is co-expressed with UFO (or UFO-VP16) but not by either effector alone (Figure 1A and 1E). We also tested the promoter of RABBIT EARS ( RBE ), a gene expressed in petal primordia in a UFO-dependent manner (Krizek et al, 2006).…”

“…Hence, in protoplasts like in plants (Lamb et al, 2002), pAP3 LFYBS at DEE is not sufficient to explain pAP3 activation. Previous studies suggested that LFY-UFO activation might involve the PEE and the region directly upstream (Hill et al, 1998; Lamb et al, 2002; Tilly et al, 1998). Indeed, in the protoplasts assay, we found that deleting the 107-bp region upstream of the PEE reduced LFY-UFO-dependent pAP3 activation (Figure 2B).…”

“…Thus, we searched for the pAP3 element(s) required for such activation. Several regulatory regions driving AP3 expression in early floral meristem have been identified, including the Distal Early Element (DEE) and the Proximal Early Element (PEE; Figure 2A; Hill et al, 1998). The DEE contains a predicted canonical LFYBS (Lamb et al, 2002; Figure S2A).…”

Functional identification of the different regions in B‐class floral homeotic MADS‐box proteins IiAP3 and IiPI from Isatis indigotica

“…A series of ORFs, including IiAP3 , IiPI and chimeras of IiAP3 and IiPI , were prepared by PCR or overlapping PCR, and were inserted into the binary expression vectors pCAMBIA 1302 possessing a GFP gene controlled by CaMV 35S promoter (Benfey & Chua, 1990), and pRI 101‐AN containing CaMV 35S promoter or 5D2 promoter, to drive a constitutive or tissue‐specific expression, respectively, of the transgenes in wild‐type Arabidopsis plants or ap3‐6 and pi‐1 mutants. The 1036 bp 5D2 promoter of AtAP3 was used as a tissue‐specific regulatory sequence in petals and stamens (Hill et al, 1998). The final constructs were verified by restriction digestion, PCR and DNA sequencing.…”