BackgroundAuxin signaling is vital for plant growth and development, and plays important role in apical dominance, tropic response, lateral root formation, vascular differentiation, embryo patterning and shoot elongation. Auxin Response Factors (ARFs) are the transcription factors that regulate the expression of auxin responsive genes. The ARF genes are represented by a large multigene family in plants. The first draft of full maize genome assembly has recently been released, however, to our knowledge, the ARF gene family from maize (ZmARF genes) has not been characterized in detail.ResultsIn this study, 31 maize (Zea mays L.) genes that encode ARF proteins were identified in maize genome. It was shown that maize ARF genes fall into related sister pairs and chromosomal mapping revealed that duplication of ZmARFs was associated with the chromosomal block duplications. As expected, duplication of some ZmARFs showed a conserved intron/exon structure, whereas some others were more divergent, suggesting the possibility of functional diversification for these genes. Out of these 31 ZmARF genes, 14 possess auxin-responsive element in their promoter region, among which 7 appear to show small or negligible response to exogenous auxin. The 18 ZmARF genes were predicted to be the potential targets of small RNAs. Transgenic analysis revealed that increased miR167 level could cause degradation of transcripts of six potential targets (ZmARF3, 9, 16, 18, 22 and 30). The expressions of maize ARF genes are responsive to exogenous auxin treatment. Dynamic expression patterns of ZmARF genes were observed in different stages of embryo development.ConclusionsMaize ARF gene family is expanded (31 genes) as compared to Arabidopsis (23 genes) and rice (25 genes). The expression of these genes in maize is regulated by auxin and small RNAs. Dynamic expression patterns of ZmARF genes in embryo at different stages were detected which suggest that maize ARF genes may be involved in seed development and germination.

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The Asian soybean rust fungus, Phakopsora pachyrhizi, is an obligate biotrophic pathogen causing severe soybean disease epidemics. Molecular mechanisms by which P. pachyrhizi and other rust fungi interact with their host plants are poorly understood. The genomes of all rust fungi encode many small, secreted cysteine-rich proteins (SSCRP). While these proteins are thought to function within the host, their roles are completely unknown. Here, we present the characterization of P. pachyrhizi effector candidate 23 (PpEC23), a SSCRP that we show to suppress plant immunity. Furthermore, we show that PpEC23 interacts with soybean transcription factor GmSPL12l and that soybean plants in which GmSPL12l is silenced have constitutively active immunity, thereby identifying GmSPL12l as a negative regulator of soybean defenses. Collectively, our data present evidence for a virulence function of a rust SSCRP and suggest that PpEC23 is able to suppress soybean immune responses and physically interact with soybean transcription factor GmSPL12l, a negative immune regulator.

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In Arabidopsis (Arabidopsis thaliana) plants growing under normal conditions, DEHYDRATION-RESPONSIVE ELEMENT BINDING PROTEIN2A (DREB2A) is present at low levels because it is ubiquitinated and destabilized by DREB2A INTERACTING PROTEIN1 (DRIP1) and DRIP2 through 26S proteasome-mediated proteolysis. Drought stress counteracts the ubiquitination and proteolysis of DREB2A, thus allowing the accumulation of sufficient amounts of DREB2A protein to activate downstream gene expression. The mechanisms leading to drought stress-mediated DREB2A accumulation are still unclear. Here, we report that the wheat (Triticum aestivum) TaSAP5 protein, which contains an A20/AN1 domain, acts as an E3 ubiquitin ligase to mediate DRIP degradation and thus increase DREB2A protein levels. Drought induces TaSAP5 expression in wheat, and TaSAP5 overexpression in Arabidopsis and wheat seedlings increased their drought tolerance, as measured by survival rate and grain yield under severe drought stress. TaSAP5 can interact with and ubiquitinate TaDRIP, as well as AtDRIP1 and AtDRIP2, leading to their subsequent degradation through the 26S proteasome pathway. Consistent with this, TaSAP5 overexpression enhances DRIP degradation and increases the levels of DREB2A protein and its downstream targets. These results suggest that TaSAP5 acts to link drought with DREB2A accumulation and illustrate the molecular mechanisms involved in this process.

The soil borne fungus, Fusarium virguliforme, causes sudden death syndrome (SDS) in soybean, which is a serious foliar and root rot disease. The pathogen has never been isolated from the diseased foliar tissues; phytotoxins produced by the pathogen are believed to cause foliar SDS symptoms. One of these toxins, a 13.5-kDa acidic protein named FvTox1, has been hypothesized to interfere with photosynthesis in infected soybean plants and cause foliar SDS. The objective of this study is to determine if FvTox1 is involved in foliar SDS development. We created and studied five independent knockout fvtox1 mutants to study the function of FvTox1. We conducted Agrobacterium tumefaciens-mediated transformation to accomplish homologous recombination of FvTox1 with a hygromycin B resistance gene, hph, to generate the fvtox1 mutants. Approximately 40 hygromycin-resistant transformants were obtained from 10(6) conidial spores of the F. virguliforme Mont-1 isolate when the spores were co-cultivated with the A. tumefaciens EHA105 but not with LBA4044 strain carrying a recombinant binary plasmid, in which the hph gene encoding hygromycin resistance was flanked by 5'- and 3'-end FvTox1 sequences. We observed homologous recombination-mediated integration of hph into the FvTox1 locus among five independent fvtox1 mutants. In stem-cutting assays using cut soybean seedlings fed with cell-free F. virguliforme culture filtrates, the knockout fvtox1 mutants caused chlorophyll losses and foliar SDS symptoms, which were over twofold less than those caused by the virulent F. virguliforme Mont-1 isolate. Similarly, in root inoculation assays, more than a twofold reduction in foliar SDS development and chlorophyll losses was observed among the seedlings infected with the fvtox1 mutants as compared to the seedlings infected with the wild-type Mont-1 isolate. These results suggest that FvTox1 is a major virulence factor involved in foliar SDS development in soybean. It is expected that interference of the function of this toxin in transgenic soybean plants will lead to generation of SDS-resistant soybean cultivars.

In soybean, the W4 gene encoding dihydroflavonol-4-reductase controls anthocyanin pigment biosynthesis in flowers. The mutant allele, w4-m, is characterized by variegated flowers and was evolved from the insertion of an endogenous transposable element, Tgm9, in intron II of the W4 gene. In the w4-m mutant line, reversion of the unstable allele from variegated to normal purple flower in revertants would indicate Tgm9's excision accompanied by its insertion into a second locus. We identified a male-sterile, female-sterile mutant from such germinal revertant bearing purple flowers. The objectives of our investigation were to map the sterility locus, identify candidate genes for the male-fertile, female-fertile phenotype, and then determine if sterility is associated with the insertion of Tgm9 in the sterility locus. We used bulked segregant analysis to map the locus to molecular linkage group J (chromosome 16). Fine mapping enabled us to flank the locus to a 62-kb region that contains only five predicted genes. One of the genes in that region, Glyma16g07850.1, codes for a helicase. A rice homolog of this gene has been shown to control crossing over and fertility phenotype. Thus, Glyma16g07850.1 is most likely the gene regulating the male and female fertility phenotype in soybean. DNA blot analysis of the segregating individuals for Tgm9 showed perfect association between sterility and the presence of the transposon. Most likely, the sterility mutation was caused by the insertion of Tgm9. The transposable element should facilitate identification of the male- and female-fertility gene. Characterization of the fertility gene will provide vital molecular insight on the reproductive biology of soybean and other plants.

The W4 locus in soybean encodes a dihydroflavonol-4-reductase (DFR2) that regulates pigmentation patterns in flowers and hypocotyl. The mutable w4-m allele that governs variegated flowers has arisen through insertion of a CACTA-type transposable element, Tgm9, in DFR2. In the w4-m line, reversion from variegated to purple flower indicates excision of Tgm9, and its insertion at a new locus. Previously, we have identified a male-sterile, female-sterile mutant among the selfed progenies of a revertant plant carrying only purple flowers. Co-segregation between Tgm9 and the sterility phenotype suggested that the mutant was generated by insertion of Tgm9 at the St8 locus. The transposon was localized to exon 10 of Glyma.16G072300 that shows high identity to the MER3 DNA helicase involved in crossing over. Molecular analysis of fertile branches from two independent revertant plants confirmed precise excision of Tgm9 from the st8 allele, which restored fertility. In soybean, the gene is expressed in flower-buds, trifoliate leaves and stem. Phylogenetic analysis placed St8 in a clade with the Arabidopsis and rice MER3 suggesting that St8 is most likely the orthologous MER3 soybean gene. This study established the utility of Tgm9 in gene identification as well as in forward and reverse genetics studies.