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JXB Advance Access originally published online on September 27, 2006
Journal of Experimental Botany 2006 57(13):3419-3431; doi:10.1093/jxb/erl144
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© The Author [2006]. Published by Oxford University Press [on behalf of the Society for Experimental Biology]. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org

Flower Development

Protein interactions of MADS box transcription factors involved in flowering in Lolium perenne

Stefano Ciannamea1, Kerstin Kaufmann1, Marta Frau1, Isabella A. Nougalli Tonaco1, Klaus Petersen2, Klaus K. Nielsen3, Gerco C. Angenent1 and Richard G. H. Immink1,*

1Plant Research International, Business unit Bioscience, Bornsesteeg 65, 6708 PD, Wageningen, The Netherlands
2Risoe National Laboratory, Biosystems Department 330, PO Box 49, 4000 Roskilde, Denmark
3DLF-TRIFOLIUM A/S 31, Hoejerupvej, PO Box 19, DK-4660 Store Heddinge, Denmark

* To whom correspondence should be addressed. E-mail: Richard.Immink{at}wur.nl

Regulation of flowering time is best understood in the dicot model species Arabidopsis thaliana. Molecular analyses revealed that genes belonging to the MADS box transcription factor family play pivotal regulatory roles in both the vernalization- and photoperiod-regulated flowering pathways. Here the analysis of three APETALA1 (AP1)-like MADS box proteins (LpMADS1–3) and a SHORT VEGETATIVE PHASE (SVP)-like MADS box protein (LpMADS10) from the monocot perennial grass species Lolium perenne is reported. Features of these MADS box proteins were studied by yeast two-hybrid assays. Protein–protein interactions among the Lolium proteins and with members of the Arabidopsis MADS box family have been studied. The expression pattern for LpMADS1 and the protein properties suggest that not the Arabidopsis AP1 gene, but the SUPPRESSOR OF CONSTANS1 (SOC1) gene, is the functional equivalent of LpMADS1. To obtain insight into the molecular mechanism underlying the regulation of LpMADS1 gene expression in vernalization-sensitive and -insensitive Lolium accessions, the upstream sequences of this gene from a winter and spring growth habit variety were compared with respect to MADS box protein binding. In both promoter elements, a putative MADS box transcription factor-binding site (CArG-box) is present; however, the putative spring promoter has a short deletion adjacent to this DNA motif. Experiments using yeast one-hybrid and gel retardation assays demonstrated that the promoter element is bound by an LpMADS1–LpMADS10 higher order protein complex and, furthermore, that this complex binds efficiently to the promoter element from the winter variety only. This strongly supports the model that LpMADS1 together with LpMADS10 controls the vernalization-dependent regulation of the LpMADS1 gene, which is part of the vernalization-induced flowering process in Lolium.

Key words: Lolium, MADS box transcription factor, protein–protein interaction, vernalization


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