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Bacterial xenogeneic silencing proteins selectively bind to and silence expression from many AT rich regions of the chromosome. They serve as master regulators of horizontally acquired DNA, including a large number of virulence genes. To date, three distinct families of xenogeneic silencers have been identified: H-NS of Proteobacteria, Lsr2 of the Actinomycetes, and MvaT of Pseudomonas sp. Although H-NS and Lsr2 family proteins are structurally different, they all recognize the AT-rich DNA minor groove through a common AT-hook-like motif, which is absent in the MvaT family. Thus, the DNA binding mechanism of MvaT has not been determined. Here, we report the characteristics of DNA sequences targeted by MvaT with protein binding microarrays, which indicates that MvaT prefers binding flexible DNA sequences with multiple TpA steps. We demonstrate that there are clear differences in sequence preferences between MvaT and the other two xenogeneic silencer families. We also determined the structure of the DNA-binding domain of MvaT in complex with a high affinity DNA dodecamer using solution NMR. This is the first experimental structure of a xenogeneic silencer in complex with DNA, which reveals that MvaT recognizes the AT-rich DNA both through base readout by an “AT-pincer” motif inserted into the minor groove and through shape readout by multiple lysine side chains interacting with the DNA sugar-phosphate backbone. Mutations of key MvaT residues for DNA binding confirm their importance with both in vitro and in vivo assays. This novel DNA binding mode enables MvaT to better tolerate GC-base pair interruptions in the binding site and less prefer A tract DNA when compared to H-NS and Lsr2. Comparison of MvaT with other bacterial xenogeneic silencers provides a clear picture that nature has evolved unique solutions for different bacterial genera to distinguish foreign from self DNA.

Summary Family 1 glycosyltransferases comprise the greatest number of glycosyltransferases found in plants. The widespread occurrence and diversity of glycosides throughout the plant kingdom underscore the importance of these glycosyltransferases. Here, we describe the identification and characterization of a late‐flowering Arabidopsis (Arabidopsis thaliana) mutant, in which a putative family 1 glycosyltransferase gene, UGT87A2, was disrupted. The role and possible mechanism of UGT87A2 in the regulation of flowering were analyzed by molecular, genetic and cellular approaches. The ugt87a2 mutant exhibited late flowering in both long and short days, and its flowering was promoted by vernalization and gibberellin. Furthermore, the mutant flowering phenotype was rescued by the wild‐type UGT87A2 gene in complementation lines. Interestingly, the expression of the flowering repressor FLOWERING LOCUS C was increased substantially in the mutant, but decreased to the wild‐type level in complementation lines, with corresponding changes in the expression levels of the floral integrators and floral meristem identity genes. The expression of UGT87A2 was developmentally regulated and its protein products were distributed in both cytoplasm and nucleus. Our findings imply that UGT87A2 regulates flowering time via the flowering repressor FLOWERING LOCUS C. These data highlight an important role for the family 1 glycosyltransferases in the regulation of plant flower development.

Trans-zeatin is a kind of cytokinins that plays a crucial role in plant growth and development. The master trans-zeatin O-glucosyltransferase of Arabidopsis thaliana, UGT85A1, has been previously identified through biochemical approach. To determine the in planta role of UGT85A1 gene, the characterization of transgenic Arabidopsis plants overexpressing UGT85A1 was carried out. Under normal conditions, transgenic Arabidopsis did not display clearly altered phenotypes. A remarkable alteration is that the accumulation level of the trans-zeatin O-glucosides was significantly increased in UGT85A1 overexpressing transgenic Arabidopsis, while other forms of cytokinins kept the similar concentrations compared to the wild type. When treated with exogenously applied trans-zeatin, UGT85A1 overexpressing Arabidopsis showed much less sensitivity to trans-zeatin in primary root elongation and lateral root formation. Meanwhile, the chlorophyll content of detached leaves of transgenic Arabidopsis was much lower than wild type. Studies of spatial-temporal expression patterns showed that UGT85A1 was mainly expressed in the early seedlings and developing seeds. Analysis of subcellular localization suggested that UGT85A1 was localized to cytoplasm and nucleus. Taken together, our data suggest that overexpression of Arabidopsis glucosyltransferase UGT85A1 influences trans-zeatin homeostasis and trans-zeatin responses likely through O-glucosylation in planta.