Seeds of the Arabidopsis thaliana transparent testa 1 mutant (tt1) appear yellow, due to the lack of condensed tannin pigments in the seed coat. The TT1 gene was isolated by reverse genetics using an En-1 transposon mutagenized A. thaliana population. TT1 gene expression was detected in developing ovules and young seeds only, and the gene was shown to encode a nuclear protein. Mutant seeds displayed altered morphology of the seed endothelium in which brown tannin pigments accumulate in wild-type plants, indicating that TT1 is involved in the differentiation of this cell layer. When overexpressed in transgenic A. thaliana plants, TT1 caused aberrant development and organ morphology. The protein contains a novel combination of two TFIIIA-type zinc finger motifs. Closely related motifs were detected in a number of putative proteins deduced from plant genomic and EST sequences. The new protein domain containing this type of zinc finger motifs was designated WIP, according to three strictly conserved amino acid residues. Our data indicate the existence of a small gene family in A. thaliana which is defined by the occurrence of the WIP domain. WIP genes may play important roles in regulating developmental processes, including the control of endothelium differentiation.

Chromatin-based silencing provides a crucial mechanism for the regulation of gene expression. We have identified a WD40 domain cyclophilin, CYCLOPHILIN71 (CYP71), which functions in gene repression and organogenesis in Arabidopsis thaliana. Disruption of CYP71 resulted in ectopic activation of homeotic genes that regulate meristem development. The cyp71 mutant plants displayed dramatic defects, including reduced apical meristem activity, delayed and abnormal lateral organ formation, and arrested root growth. CYP71 was associated with the chromatin of target gene loci and physically interacted with histone H3. The cyp71 mutant showed reduced methylation of H3K27 at target loci, consistent with the derepression of these genes in the mutant. As CYP71 has close homologs in eukaryotes ranging from fission yeast to human, we propose that it serves as a highly conserved histone remodeling factor involved in chromatin-based gene silencing in eukaryotic organisms.

SUMMARYWild type seed coats of Arabidopsis thaliana are brown due to the accumulation of proanthocyanidin pigments (PAs). The pigmentation requires activation of phenylpropanoid biosynthesis genes and mutations in some of these genes cause a yellow appearance of seeds, termed transparent testa (tt) phenotype. The TT1 gene encodes a WIP-type zinc finger protein and is expressed in the seed coat endothelium where most of the PAs accumulate in wild type plants. In this study we show that TT1 is not only required for correct expression of PA-specific genes in the seed coat, but also affects CHS, encoding the first enzyme of flavonoid biosynthesis. Many steps of this pathway are controlled by complexes of MYB and BHLH proteins with the WD40 factor TTG1. We demonstrate that TT1 can interact with the R2R3 MYB protein TT2 and that ectopic expression of TT2 can partially restore the lack in PA production in tt1. Reduced seed coat pigmentation was obtained using a TT1 variant lacking nuclear localisation signals. Based on our results we propose that the TT2/TT8/TTG1 regulon may also comprise early genes like CHS and discuss steps to further unravel the regulatory network controlling flavonoid accumulation in endothelium cells during A. thaliana seed development.

a b s t r a c t WIP proteins form a plant specific subfamily of C2H2 zinc finger (ZF) proteins. In this study, we functionally characterized the WIP domain, which consists of four ZF motifs, and discuss molecular functions for WIP proteins. Mutations in each of the ZFs lead to loss of function of the TT1/WIP1 protein in Arabiopsis thaliana. SV40 type nuclear localisation signals were detected in two of the ZFs and functionally characterized using GFP fusions as well as new mutant alleles identified by TILLING. Promoter swap experiments showed that selected WIP proteins are partially able to take over TT1 function. Activity of the AtBAN promoter, a potential TT1 target, could be increased by the addition of TT1 to the TT2-TT8-TTG1 regulatory complex.

The global commercial cultivation of transgenic crops, including glyphosate-tolerant soybean, has increased widely in recent decades with potential impact on the environment. The bulk of previous studies showed different results on the effects of the release of transgenic plants on the soil microbial community, especially rhizosphere bacteria. In this study, comparative analyses of the bacterial communities in the rhizosphere soils and surrounding soils were performed between the glyphosate-tolerant soybean line NZL06-698 (or simply N698), containing a glyphosate-insensitive gene, and its control cultivar Mengdou12 (or simply MD12), by a 16S ribosomal RNA gene (16S rDNA) amplicon sequencing-based Illumina MiSeq platform. No statistically significant difference was found in the overall alpha diversity of the rhizosphere bacterial communities, although the species richness and evenness of the bacteria increased in the rhizosphere of N698 compared with that of MD12. Some influence on phylogenetic diversity of the rhizosphere bacterial communities was found between N698 and MD12 by beta diversity analysis based on weighted UniFrac distance. Furthermore, the relative abundances of part rhizosphere bacterial phyla and genera, which included some nitrogen-fixing bacteria, were significantly different between N698 and MD12. Our present results indicate some impact of the glyphosate-tolerant soybean line N698 on the phylogenetic diversity of rhizosphere bacterial communities together with a significant difference in the relative abundances of part rhizosphere bacteria at different classification levels as compared with its control cultivar MD12, when a comparative analysis of surrounding soils between N698 and MD12 was used as a systematic contrast study.