SummaryIn-depth analysis of protein-protein interaction specificities of the MYB protein family of Arabidopsis thaliana revealed a conserved amino acid signature ([DE]Lx 2 [RK]x 3 Lx 6 Lx 3 R) as the structural basis for interaction between MYB and R/B-like BHLH proteins. The motif has successfully been used to predict new MYB/BHLH interactions for A. thaliana proteins, it allows to discriminate between even closely related MYB proteins and it is conserved amongst higher plants. In A. thaliana, the motif is shared by fourteen R2R3 MYB proteins and six 1R MYB proteins. It is located on helices 1 and 2 of the R3 repeat and forms a characteristic surface-exposed pattern of hydrophobic and charged residues. Single-site mutation of any amino acid of the signature impairs the interaction. Two particular amino acids have been determined to account for most of the interaction stability. Functional specificity of MYB/BHLH complexes was investigated in vivo by a transient DFR promoter activation assay. Residues stabilizing the MYB/BHLH interaction were shown to be critical for promoter activation. By virtue of proved and predicted interaction specificities, this study provides a comprehensive survey of the MYB proteins that interact with R/B-like BHLH proteins potentially involved in the TTG1-dependent regulatory interaction network. The results are discussed with respect to multi-functionality, specificity and redundancy of MYB and BHLH protein function.

The organization of living cells is based on networks of interacting molecules. Systematic analysis of protein interactions of 3-aa loop extension (TALE) homeodomain proteins, fundamental regulators of plant meristem function and leaf development, revealed a highly connected, complex network. The network includes nine members of Arabidopsis thaliana ovate family proteins (AtOFPs), a plant-specific protein family, indicating a close functional connection to TALE homeodomain proteins. Evidence is provided that AtOFP1 is an essential pleiotropic developmental regulator. At-OFP1 and AtOFP5 are shown to associate with the cytoskeleton and to regulate subcellular localization of TALE homeodomain proteins, suggesting a previously unrecognized control mechanism in plant development.cytoskeleton ͉ network modules ͉ protein-protein interactions ͉ BELL proteins ͉ KNOX proteins

Mitogen-activated protein kinase (MAPK)-mediated responses are in part regulated by the repertoire of MAPK substrates, which is still poorly elucidated in plants. Here, the in vivo enzyme-substrate interaction of the Arabidopsis thaliana MAP kinase, MPK6, with an ethylene response factor (ERF104) is shown by fluorescence resonance energy transfer. The interaction was rapidly lost in response to flagellin-derived flg22 peptide. This complex disruption requires not only MPK6 activity, which also affects ERF104 stability via phosphorylation, but also ethylene signaling. The latter points to a novel role of ethylene in substrate release, presumably allowing the liberated ERF104 to access target genes. Microarray data show enrichment of GCC motifs in the promoters of ERF104 -up-regulated genes, many of which are stress related. ERF104 is a vital regulator of basal immunity, as altered expression in both erf104 and overexpressors led to more growth inhibition by flg22 and enhanced susceptibility to a non-adapted bacterial pathogen.itogen-activated protein kinase (MAPK) cascades transduce external signals into cellular responses in eukaryotes (1). In plants, MAPKs orthologous to the Arabidopsis MPK3, MPK4, and MPK6 are activated by various stimuli including flg22, a bacterial flagellin-derived peptide that acts as a pathogen-associated molecular pattern (PAMP) (2-5). These three MAPKs control defense positively (MPK3/MPK6) (3, 6) or negatively (MPK4) (7).Many phytohormones have been shown to affect defense responses; but most progress has been made in regard to salicylic acid (SA), jasmonic acid (JA) and ethylene (ET) (8). The tobacco MPK6 ortholog is activated by SA (9) and the Arabidopsis mpk4 mutant has elevated SA levels and enhanced pathogen resistance (7). Genetic evidence linking ET to MAPK signaling is also suggested by the negative regulator of the ET response, Constitutive Triple Response 1 (CTR1), a Raf-like kinase that was recently shown to control MPK3/6 activation via MKK9 (MAPK kinase 9) (10). Both JA and the ET precursor, 1-aminocyclopropane-1-carboxylic acid (ACC), activate MPK6 in Arabidopsis (11, 12) but not in tobacco (13). Although responses may differ between plant species, the activation of MPK6 by ET/ACC is highly debated (14). In another report, ACC did not activate MPK6, but ET biosynthesis was positively regulated by MPK6 through posttranslational stabilization of the rate-limiting ACC synthase (ACS) isoforms, ACS2 and ACS6 (14,15).In addition to the cytoplasmic ACSs, MAPKs also target nuclear proteins (10,16,17); this may occur either after MAPK nuclear translocation following activation (18,19) or as preformed nuclear protein complexes (20). The latter would imply movement of the upstream MKKs into the nucleus to modify the MAPKs or, alternatively, that the activated MAPKs enter the nucleus to displace the inactive MAPK from preformed complexes. Examples of nuclear targets include the MPK4 substrates, MKS1 and two MKS1-interactors of the WRKY transcription factor family, WRKY25 and WRKY33 ...

SUMMARYAnthocyanins are natural pigments that accumulate only in light-grown and not in dark-grown Arabidopsis plants. Repression of anthocyanin accumulation in darkness requires the CONSTITUTIVELY PHOTOMORPH-OGENIC1/SUPPRESSOR OF PHYA-105 (COP1/SPA) ubiquitin ligase, as cop1 and spa mutants produce anthocyanins also in the dark. Here, we show that COP1 and SPA proteins interact with the myeloblastosis (MYB) transcription factors PRODUCTION OF ANTHOCYANIN PIGMENT1 (PAP)1 and PAP2, two members of a small protein family that is required for anthocyanin accumulation and for the expression of structural genes in the anthocyanin biosynthesis pathway. The increased anthocyanin levels in cop1 mutants requires the PAP1 gene family, indicating that COP1 functions upstream of the PAP1 gene family. PAP1 and PAP2 proteins are degraded in the dark and this degradation is dependent on the proteasome and on COP1. Hence, the light requirement for anthocyanin biosynthesis results, at least in part, from the light-mediated stabilization of PAP1 and PAP2. Consistent with this conclusion, moderate overexpression of PAP1 leads to an increase in anthocyanin levels only in the light and not in darkness. Here we show that SPA genes are also required for reducing PAP1 and PAP2 transcript levels in dark-grown seedlings. Taken together, these results indicate that the COP1/SPA complex affects PAP1 and PAP2 both transcriptionally and post-translationally. Thus, our findings have identified mechanisms via which the COP1/SPA complex controls anthocyanin levels in Arabidopsis that may be useful for applications in biotechnology directed towards increasing anthocyanin content in plants.