DNA-binding properties of the MADS-domain transcription factor SEPALLATA3 and mutant variants characterized by SELEX-seq

Key message We studied the DNA-binding profile of the MADS-domain transcription factor SEPALLATA3 and mutant variants by SELEX-seq. DNA-binding characteristics of SEPALLATA3 mutant proteins lead us to propose a novel DNA-binding mode. MIKC-type MADS-domain proteins, which function as essential transcription factors in plant development, bind as dimers to a 10-base-pair AT-rich motif termed CArG-box. However, this consensus motif cannot fully explain how the abundant family members in flowering plants can bind different target genes in specific ways. The aim of this study was to better understand the DNA-binding specificity of MADS-domain transcription factors. Also, we wanted to understand the role of a highly conserved arginine residue for binding specificity of the MADS-domain transcription factor family. Here, we studied the DNA-binding profile of the floral homeotic MADS-domain protein SEPALLATA3 by performing SELEX followed by high-throughput sequencing (SELEX-seq). We found a diverse set of bound sequences and could estimate the in vitro binding affinities of SEPALLATA3 to a huge number of different sequences. We found evidence for the preference of AT-rich motifs as flanking sequences. Whereas different CArG-boxes can act as SEPALLATA3 binding sites, our findings suggest that the preferred flanking motifs are almost always the same and thus mostly independent of the identity of the central CArG-box motif. Analysis of SEPALLATA3 proteins with a single amino acid substitution at position 3 of the DNA-binding MADS-domain further revealed that the conserved arginine residue, which has been shown to be involved in a shape readout mechanism, is especially important for the recognition of nucleotides at positions 3 and 8 of the CArG-box motif. This leads us to propose a novel DNA-binding mode for SEPALLATA3, which is different from that of other MADS-domain proteins known.Peer reviewe

Multi-Jet Event Rates in Deep Inelastic Scattering and Determination of the Strong Coupling Constant

Jet event rates in deep inelastic ep scattering at HERA are investigated applying the modified JADE jet algorithm. The analysis uses data taken with the H1 detector in 1994 and 1995. The data are corrected for detector and hadronization effects and then compared with perturbative QCD predictions using next-to-leading order calculations. The strong coupling constant alpha_S(M_Z^2) is determined evaluating the jet event rates. Values of alpha_S(Q^2) are extracted in four different bins of the negative squared momentum transfer~\qq in the range from 40 GeV2 to 4000 GeV2. A combined fit of the renormalization group equation to these several alpha_S(Q^2) values results in alpha_S(M_Z^2) = 0.117+-0.003(stat)+0.009-0.013(syst)+0.006(jet algorithm).Comment: 17 pages, 4 figures, 3 tables, this version to appear in Eur. Phys. J.; it replaces first posted hep-ex/9807019 which had incorrect figure 4

Measurement of Leading Proton and Neutron Production in Deep Inelastic Scattering at HERA

Deep--inelastic scattering events with a leading baryon have been detected by the H1 experiment at HERA using a forward proton spectrometer and a forward neutron calorimeter. Semi--inclusive cross sections have been measured in the kinematic region 2 <= Q^2 <= 50 GeV^2, 6.10^-5 <= x <= 6.10^-3 and baryon p_T <= MeV, for events with a final state proton with energy 580 <= E' <= 740 GeV, or a neutron with energy E' >= 160 GeV. The measurements are used to test production models and factorization hypotheses. A Regge model of leading baryon production which consists of pion, pomeron and secondary reggeon exchanges gives an acceptable description of both semi-inclusive cross sections in the region 0.7 <= E'/E_p <= 0.9, where E_p is the proton beam energy. The leading neutron data are used to estimate for the first time the structure function of the pion at small Bjorken--x.Comment: 30 pages, 9 figures, 2 tables, submitted to Eur. Phys.

Dynamic genome evolution in a model fern

The large size and complexity of most fern genomes have hampered efforts to elucidate fundamental aspects of fern biology and land plant evolution through genome-enabled research. Here we present a chromosomal genome assembly and associated methylome, transcriptome and metabolome analyses for the model fern species Ceratopteris richardii. The assembly reveals a history of remarkably dynamic genome evolution including rapid changes in genome content and structure following the most recent whole-genome duplication approximately 60 million years ago. These changes include massive gene loss, rampant tandem duplications and multiple horizontal gene transfers from bacteria, contributing to the diversification of defence-related gene families. The insertion of transposable elements into introns has led to the large size of the Ceratopteris genome and to exceptionally long genes relative to other plants. Gene family analyses indicate that genes directing seed development were co-opted from those controlling the development of fern sporangia, providing insights into seed plant evolution. Our findings and annotated genome assembly extend the utility of Ceratopteris as a model for investigating and teaching plant biology

Jets and energy flow in photon-proton collisions at HERA

Properties of the hadronic final state in photoproduction events with large transverse energy are studied at the electron-proton collider HERA. Distributions of the transverse energy, jets and underlying event energy are compared to \overline{p}p data and QCD calculations. The comparisons show that the \gamma p events can be consistently described by QCD models including -- in addition to the primary hard scattering process -- interactions between the two beam remnants. The differential jet cross sections d\sigma/dE_T^{jet} and d\sigma/d\eta^{jet} are measured

A hitchhiker's guide to the MADS world of plants

Plant life critically depends on the function of MADS-box genes encoding MADS-domain transcription factors, which are present to a limited extent in nearly all major eukaryotic groups, but constitute a large gene family in land plants. There are two types of MADS-box genes, termed type I and type II, and in plants these groups are distinguished by exon-intron and domain structure, rates of evolution, developmental function and degree of functional redundancy. The type I genes are further subdivided into three groups - Mα, Mβ and Mγ - while the type II genes are subdivided into the MIKC(C )and MIKC* groups. The functional diversification of MIKC(C )genes is closely linked to the origin of developmental and morphological novelties in the sporophytic (usually diploid) generation of seed plants, most spectacularly the floral organs and fruits of angiosperms. Functional studies suggest different specializations for the different classes of genes; whereas type I genes may preferentially contribute to female gametophyte, embryo and seed development and MIKC*-group genes to male gametophyte development, the MIKC(C)-group genes became essential for diverse aspects of sporophyte development. Beyond the usual transcriptional regulation, including feedback and feed-forward loops, various specialized mechanisms have evolved to control the expression of MADS-box genes, such as epigenetic control and regulation by small RNAs. In future, more data from genome projects and reverse genetic studies will allow us to understand the birth, functional diversification and death of members of this dynamic and important family of transcription factors in much more detail

Promoter analysis of MADS-box genes in eudicots through phylogenetic footprinting

The MIKC MADS-box gene family has been shaped by extensive gene duplications giving rise to subfamilies of genes with distinct functions and expression patterns. However, within these subfamilies the functional assignment is not that clear-cut, and considerable functional redundancy exists. One way to investigate the diversity in regulation present in these subfamilies is promoter sequence analysis. With the advent of genome sequencing projects, we are now able to exert a comparative analysis of Arabidopsis and poplar promoters of MADS-box genes belonging to the same subfamily. Based on the principle of phylogenetic footprinting, sequences conserved between the promoters of homologous genes are thought to be functional. Here, we have investigated the evolution of MADS-box genes at the promoter level and show that many genes have diverged in their regulatory sequences after duplication and/or speciation. Furthermore, using phylogenetic footprinting, a distinction can be made between redundancy, neo/nonfunctionalization, and subfunctionalization

FLOWERING LOCUS C in monocots and the tandem origin of angiosperm specific MADS-box genes

MADS-domain transcription factors have been shown to act as key repressors or activators of the transition to flowering and as master regulators of reproductive organ identities. Despite their important roles in plant development, the origin of several MADS-box subfamilies has remained enigmatic so far. Here we demonstrate, through a combination of genome synteny and phylogenetic reconstructions, the origin of three major, apparently angiosperm-specific MADS-box gene clades: FLOWERING LOCUS C- (FLC-), SQUAMOSA- (SQUA-) and SEPALLATA- (SEP-)-like genes. We find that these lineages derive from a single ancestral tandem duplication in a common ancestor of extant seed plants. Contrary to common belief, we show that FLC-like genes are present in cereals where they can also act as floral repressors responsive to prolonged cold or vernalization. This opens a new perspective on the translation of findings from Arabidopsis to cereal crops, in which vernalization was originally described.status: publishe

The ABCs of flower development: mutational analysis of AP1/FUL-like genes in rice provides evidence for a homeotic (A)-function in grasses

The well-known ABC model describes the combinatorial interaction of homeotic genes in specifying floral organ identities. While the B-and C-functions are highly conserved throughout flowering plants and even in gymnosperms, the A-function, which specifies the identity of perianth organs (sepals and petals in eudicots), remains controversial. One reason for this is that in most plants that have been investigated thus far, with Arabidopsis being a remarkable exception, one does not find recessive mutants in which the identity of both types of perianth organs is affected. Here we report a comprehensive mutational analysis of all four members of the AP1/FUL-like subfamily of MADS-box genes in rice (Oryza sativa). We demonstrate that OsMADS14 and OsMADS15, in addition to their function of specifying meristem identity, are also required to specify palea and lodicule identities. Because these two grass-specific organs are very likely homologous to sepals and petals of eudicots, respectively, we conclude that there is a floral homeotic (A)-function in rice as defined previously. Together with other recent findings, our data suggest that AP1/FUL-like genes were independently recruited to fulfil the (A)-function in grasses and some eudicots, even though other scenarios cannot be excluded and are discussed