SummaryIn Arabidopsis thaliana, several MYB and basic helix-loop-helix (BHLH) proteins form ternary complexes with TTG1 (WD-Repeats) and regulate the transcription of genes involved in anthocyanin and proanthocyanidin (PA) biosynthesis. Similar MYB-BHLH-WDR (MBW) complexes control epidermal patterning and cell fates. A family of small MYB proteins (R3-MYB) has been shown to play an important role in the regulation of epidermal cell fates, acting as inhibitors of the MBW complexes. However, so far none of these small MYB proteins have been demonstrated to regulate flavonoid biosynthesis. The genetic and molecular analyses presented here demonstrated that Arabidopsis MYBL2, which encodes a R3-MYB-related protein, is involved in the regulation of flavonoid biosynthesis. The loss of MYBL2 activity in the seedlings of two independent T-DNA insertion mutants led to a dramatic increase in the accumulation of anthocyanin. In addition, overexpression of MYBL2 in seeds inhibited the biosynthesis of PAs. These changes in flavonoid content correlate well with the increased level of mRNA of several structural and regulatory anthocyanin biosynthesis genes. Interestingly, transient expression analyses in A. thaliana cells suggested that MYBL2 interacts with MBW complexes in planta and directly modulates the expression of flavonoid target genes. These results are fully consistent with the molecular interaction of MYBL2 with BHLH proteins observed in yeast. Finally, MYBL2 expression studies, including its inhibition by light-induced stress, allowed us to hypothesise a physiological role for MYBL2. Taken together, these results bring new insights into the transcriptional regulation of flavonoid biosynthesis and provide new clues and tools for further investigation of its developmental and environmental regulation.Keywords: flavonoid, transcription, network, MYB, bHLH, TTG1. IntroductionFlavonoids are secondary metabolites that fulfil important biological functions and provide useful metabolic and genetic models for plant research, including the analysis of transcriptional regulation of gene expression (Koes et al., 2005;Lepiniec et al., 2006;Peer and Murphy, 2007;Taylor and Grotewold, 2005;Winkel-Shirley, 2001). Flavonoids are involved in protection against various biotic and abiotic stresses, they play roles in the regulation of plant reproduction and development and act as signalling molecules with the biotic environment. Besides these physiological functions, there is a growing interest in these secondary metabolites due to their potential benefits for human health (Halliwell, 2007; Luceri et al., 2007); therefore, improving our understanding of the regulation of flavonoid biosynthesis is an important objective.Although structural genes can be efficiently targeted for crop improvement, the use of regulatory genes seems to be at least as promising (Bovy et al., 2007; Grotewold et al., 940 ª 2008 The Authors Journal compilation ª 2008 Blackwell Publishing LtdThe Plant Journal (2008Journal ( ) 55, 940-953 doi: 10.1111Journal...

Nitrate is an important nitrogen source for plants, but also a signal molecule that controls various aspects of plant development. In the present study the role of nitrate on seed dormancy in Arabidopsis was investigated. The effects of either mutations affecting the Arabidopsis nitrate reductase genes or of different nitrate regimes of mother plants on the dormancy of the seeds produced were analysed. Altogether, data show that conditions favouring nitrate accumulation in mother plants and in seeds lead to a lower dormancy of seeds with little other morphological or biochemical differences. Analysis of germination during seed development indicated that nitrate does not prevent the onset of dormancy but rather its maintenance. The effect of an exogenous supply of nitrate on seed germination was tested: nitrate in contrast to glutamine or potassium chloride clearly stimulated the germination of dormant seeds. Data show, moreover, that the Arabidopsis dual affinity nitrate transporter NRT1.1 ( CHL1 ) may be involved in conveying the nitrate signal into seeds. Thus, nitrate provided exogenously or by mother plants to the produced seeds, acts as a signal molecule favouring germination in Arabidopsis . This signalling may involve interaction with the abscisic acid or gibberellin pathway.

Nitrate releases seed dormancy in Arabidopsis (Arabidopsis thaliana) Columbia accession seeds in part by reducing abscisic acid (ABA) levels. Nitrate led to lower levels of ABA in imbibed seeds when included in the germination medium (exogenous nitrate). Nitrate also reduced ABA levels in dry seeds when provided to the mother plant during seed development (endogenous nitrate). Transcript profiling of imbibed seeds treated with or without nitrate revealed that exogenous nitrate led to a higher expression of nitrate-responsive genes, whereas endogenous nitrate led to a profile similar to that of stratified or after-ripened seeds. Profiling experiments indicated that the expression of the ABA catabolic gene CYP707A2 was regulated by exogenous nitrate. The cyp707a2-1 mutant failed to reduce seed ABA levels in response to both endogenous and exogenous nitrate. In contrast, both endogenous and exogenous nitrate reduced ABA levels of the wild-type and cyp707a1-1 mutant seeds. The CYP707A2 mRNA levels in developing siliques were positively correlated with different nitrate doses applied to the mother plants. This was consistent with a role of the CYP707A2 gene in controlling seed ABA levels in response to endogenous nitrate. The cyp707a2-1 mutant was less sensitive to exogenous nitrate for breaking seed dormancy. Altogether, our data underline the central role of the CYP707A2 gene in the nitrate-mediated control of ABA levels during seed development and germination.

Photosynthetic organisms support cell metabolism by harvesting sunlight to fuel the photosynthetic electron transport. The flow of excitation energy and electrons in the photosynthetic apparatus needs to be continuously modulated to respond to dynamics of environmental conditions, and Flavodiiron (FLV) proteins are seminal components of this regulatory machinery in cyanobacteria. FLVs were lost during evolution by flowering plants, but are still present in nonvascular plants such as Physcomitrella patens. We generated P. patens mutants depleted in FLV proteins, showing their function as an electron sink downstream of photosystem I for the first seconds after a change in light intensity. flv knock-out plants showed impaired growth and photosystem I photoinhibition when exposed to fluctuating light, demonstrating FLV's biological role as a safety valve from excess electrons on illumination changes. The lack of FLVs was partially compensated for by an increased cyclic electron transport, suggesting that in flowering plants, the FLV's role was taken by other alternative electron routes.ife on Earth depends on oxygenic photosynthesis, which enables plants, algae, and cyanobacteria to convert light into chemical energy. Sunlight powers the transfer of electrons from water to NADP + by the activity of two photosystems (PS), PSII and PSI, thus generating NADPH and ATP to sustain cell metabolism. Natural environmental conditions are highly variable, and sudden changes in irradiation can drastically affect the flow of excitation energy and electrons. At the same time, the ATP and NADPH consumption rate is also highly dynamic because of a continuous metabolic regulation (1-3). Photosynthetic organisms evolved several mechanisms to modulate the flow of excitation energy and electrons according to metabolic constraints, diverting/ feeding electrons from/to the linear transport chain (3). These pathways modulate the ATP/NADPH ratio, as in the cyclic electron transport (CET) around PSI, where electrons are redirected from PSI to plastoquinone (PQ) or Cytb 6 f (4), contributing to proton translocation and ATP synthesis, but not to NADPH formation (5-9).In cyanobacteria, the Flavodiiron proteins (known as FLV) have been identified as an additional component of electron transport chain (10-12). FLV proteins are constituted by three distinct domains: a N-terminal β-lactamase-like domain, a flavodoxin-like domain, and a C-terminal NAD(P)H-flavin reductase-like domain. The former two domains are also found in FLV proteins from archaea and anaerobic bacteria, where they are involved in O 2 or NO reduction, whereas the latter is typical only of FLVs from oxygenic photosynthetic organisms (11-13). Recent studies showed that in cyanobacteria, the FLV1/FLV3 heterodimer catalyzes the light-dependent reduction of O 2 to water, using NADPH as electron donor (10, 11), protecting PSI from light stress (10). Another FLVs heterodimer, FLV2/FLV4, instead, has been shown to be active in photo-protection of PSII (14-16). FLVs also were found expre...