Aging-related changes in the diversity of women's skin microbiomes associated with oral bacteria.

Skin aging is associated with changes in cutaneous physiology including interactions with a skin microbial community. A striking alteration and diversification in the skin microbiome with aging was observed between two different age groups of 37 healthy Japanese women, i.e. younger adults of 21-37 years old and older adults of 60-76 years old, using bacterial 16S rRNA gene sequencing. The analyses revealed that the alpha diversity/species richness was significantly higher in the older than the younger group for the cheek and forehead microbiomes, while the beta diversity in the overall structure significantly differed particularly for the forearm and scalp microbiomes between the two age groups. Taxonomic profiling showed a striking reduction in the relative abundance of the majority skin genus Propionibacterium in the cheek, forearm and forehead microbiomes of the older adults, and identified 38 species including many oral bacteria that significantly differentiated the two age groups with a skin site dependency. Furthermore, we found chronological age-related and unrelated skin clinical parameters that correlate with the observed changes in the skin microbiome diversity. Thus, our data suggested that the diversification of skin microbiomes in adult women was largely affected by chronological and physiological skin aging in association with oral bacteria.

Characterization of the major bacterial-fungal populations colonizing dandruff scalps in Shanghai, China, shows microbial disequilibrium.

Dandruff is a scalp disorder characterized by the formation of flaky white-yellowish scales due to an altered proliferation and differentiation status; a disrupted barrier function; a decrease in the level of hydration and of natural moisturizing factors (NMF) in the scalp, with a persistent and relapsing inflammatory condition. It was recently reported that an imbalance between bacterial and fungal species colonizing the scalp of French volunteers was associated with dandruff condition. The purpose of the present study was to analyze the major bacterial and fungal species present on the scalp surface of Chinese volunteers and to investigate possible region-related variation in the microbiota linked to dandruff condition. The data obtained from the Chinese populations were highly similar to those obtained in France, confirming that dandruff scalps are associated with a higher incidence of Malassezia restricta and Staphylococcal sp. The ratios of Malassezia to Propionibacterium and Propionibacterium to Staphylococcus were also significantly higher in the dandruff volunteers as compared to normal volunteers, suggesting that equilibrium between the major bacterial and fungal taxa found on the normal scalps is perturbed in the dandruff scalps. The main difference between the French and Shanghai subjects was in their Staphylococcal biota. The results obtained in China and in France suggest that targeting one particular Malassezia sp. by antifungals instead of using large spectrum antifungals and rebalancing the dandruff scalp microbiota could be common approach to improve dandruff condition in the two countries.

Adventitious shoot regeneration from juvenile cotyledons of a biodiesel producing Plant, Jatropha curcas L.

An efficient and rapid adventitious shoot regeneration system from young cotyledons of Jatropha curcas L. was developed. After testing several combinations of plant growth regulators, the highest regeneration frequency was obtained for the medium supplemented with 3 mg/l BA and 0.1 mg/l IBA. This system will be useful for promoting genetic improvement efforts in J. curcas.

Sequence analysis of the genome of an oil-bearing tree, Jatropha curcas L.

The whole genome of Jatropha curcas was sequenced, using a combination of the conventional Sanger method and new-generation multiplex sequencing methods. Total length of the non-redundant sequences thus obtained was 285 858 490 bp consisting of 120 586 contigs and 29 831 singlets. They accounted for ~95% of the gene-containing regions with the average G + C content was 34.3%. A total of 40 929 complete and partial structures of protein encoding genes have been deduced. Comparison with genes of other plant species indicated that 1529 (4%) of the putative protein-encoding genes are specific to the Euphorbiaceae family. A high degree of microsynteny was observed with the genome of castor bean and, to a lesser extent, with those of soybean and Arabidopsis thaliana. In parallel with genome sequencing, cDNAs derived from leaf and callus tissues were subjected to pyrosequencing, and a total of 21 225 unigene data have been generated. Polymorphism analysis using microsatellite markers developed from the genomic sequence data obtained was performed with 12 J. curcas lines collected from various parts of the world to estimate their genetic diversity. The genomic sequence and accompanying information presented here are expected to serve as valuable resources for the acceleration of fundamental and applied research with J. curcas, especially in the fields of environment-related research such as biofuel production. Further information on the genomic sequences and DNA markers is available at http://www.kazusa.or.jp/jatropha/.

Binding of cysteine synthase to the STAS domain of sulfate transporter and its regulatory consequences.

The sulfate ion (SO(4)(2-)) is transported into plant root cells by SO(4)(2-) transporters and then mostly reduced to sulfide (S(2-)). The S(2-) is then bonded to O-acetylserine through the activity of cysteine synthase (O-acetylserine (thiol)lyase or OASTL) to form cysteine, the first organic molecule of the SO(4)(2-) assimilation pathway. Here, we show that a root plasma membrane SO(4)(2-) transporter of Arabidopsis, SULTR1;2, physically interacts with OASTL. The interaction was initially demonstrated using a yeast two-hybrid system and corroborated by both in vivo and in vitro binding assays. The domain of SULTR1;2 shown to be important for association with OASTL is called the STAS domain. This domain is at the C terminus of the transporter and extends from the plasma membrane into the cytoplasm. The functional relevance of the OASTL-STAS interaction was investigated using yeast mutant cells devoid of endogenous SO(4)(2-) uptake activity but co-expressing SULTR1;2 and OASTL. The analysis of SO(4)(2-) transport in these cells suggests that the binding of OASTL to the STAS domain in this heterologous system negatively impacts transporter activity. In contrast, the activity of purified OASTL measured in vitro was enhanced by co-incubation with the STAS domain of SULTR1;2 but not with the analogous domain of the SO(4)(2-) transporter isoform SULTR1;1, even though the SULTR1;1 STAS peptide also interacts with OASTL based on the yeast two-hybrid system and in vitro binding assays. These observations suggest a regulatory model in which interactions between SULTR1;2 and OASTL coordinate internalization of SO(4)(2-) with the energetic/metabolic state of plant root cells.

Unequal functional redundancy between the two Arabidopsis thaliana high-affinity sulphate transporters SULTR1;1 and SULTR1;2.

* In Arabidopsis, SULTR1;1 and SULTR1;2 are two genes proposed to be involved in high-affinity sulphate uptake from the soil solution. We address here the specific issue of their functional redundancy for the uptake of sulphate and for the accumulation of its toxic analogue selenate with regard to plant growth and selenate tolerance. * Using the complete set of genotypes, including the wild-type, each one of the single sultr1;1 and sultr1;2 mutants and the resulting double sultr1;1-sultr1;2 mutant, we performed a detailed phenotypic analysis of root length, shoot biomass, sulphate uptake, sulphate and selenate accumulation and selenate tolerance. * The results all ordered the four different genotypes according to the same functional hierarchy. Wild-type and sultr1;1 mutant plants displayed similar phenotypes. By contrast, sultr1;1-sultr1;2 double-mutant plants showed the most extreme phenotype and the sultr1;2 mutant displayed intermediate performances. Additionally, the degree of selenate tolerance was directly related to the seedling selenate content according to a single sigmoid regression curve common to all the genotypes. * The SULTR1;1 and SULTR1;2 genes display unequal functional redundancy, which leaves open for SULTR1;1 the possibility of displaying an additional function besides its role in sulphate membrane transport.

An Arabidopsis thaliana high-affinity molybdate transporter required for efficient uptake of molybdate from soil.

Molybdenum (Mo) is a trace element essential for living organisms, however no molybdate transporter has been identified in eukaryotes. Here, we report the identification of a molybdate transporter, MOT1, from Arabidopsis thaliana. MOT1 is expressed in both roots and shoots, and the MOT1 protein is localized, in part, to plasma membranes and to vesicles. MOT1 is required for efficient uptake and translocation of molybdate and for normal growth under conditions of limited molybdate supply. Kinetics studies in yeast revealed that the K(m) value of MOT1 for molybdate is approximately 20 nM. Furthermore, Mo uptake by MOT1 in yeast was not affected by coexistent sulfate, and MOT1 did not complement a sulfate transporter-deficient yeast mutant strain. These data confirmed that MOT1 is specific for molybdate and that the high affinity of MOT1 allows plants to obtain scarce Mo from soil.

An essential role for 14-3-3 proteins in brassinosteroid signal transduction in Arabidopsis.

Brassinosteroids (BRs) are essential hormones for plant growth and development. BRs regulate gene expression by inducing dephosphorylation of two key transcription factors, BZR1 and BZR2/BES1, through a signal transduction pathway that involves cell-surface receptors (BRI1 and BAK1) and a GSK3 kinase (BIN2). How BR-regulated phosphorylation controls the activities of BZR1/BZR2 is not fully understood. Here, we show that BIN2-catalyzed phosphorylation of BZR1/BZR2 not only inhibits DNA binding, but also promotes binding to the 14-3-3 proteins. Mutations of a BIN2-phosphorylation site in BZR1 abolish 14-3-3 binding and lead to increased nuclear localization of BZR1 protein and enhanced BR responses in transgenic plants. Further, BR deficiency increases cytoplasmic localization, and BR treatment induces rapid nuclear localization of BZR1/BZR2. Thus, 14-3-3 binding is required for efficient inhibition of phosphorylated BR transcription factors, largely through cytoplasmic retention. This study demonstrates that multiple mechanisms are required for BR regulation of gene expression and plant growth.

The role of the STAS domain in the function and biogenesis of a sulfate transporter as probed by random mutagenesis.

Sulfate transporters in plants represent a family of proteins containing transmembrane domains that constitute the catalytic part of the protein and a short linking region that joins this catalytic moiety with a C-terminal STAS domain. The STAS domain resembles an anti-sigma factor antagonist of Bacillus subtilis, which is one distinguishing feature of the SLC26 transporter family; this family includes transporters for sulfate and other anions such as iodide and carbonate. Recent work has demonstrated that this domain is critical for the activity of Arabidopsis thaliana sulfate transporters, and specific lesions in this domain, or the exchange of STAS domains between different sulfate transporters, can severely impair transport activity. In this work we generated a Saccharomyces cerevisiae expression library of the A. thaliana Sultr1;2 gene with random mutations in the linking region-STAS domain and identified STAS domain lesions that altered Sultr1;2 biogenesis and/or function. A number of mutations in the beta-sheet that forms the core of the STAS domain prevented intracellular accumulation of Sultr1;2. In contrast, the linking region and one surface of the STAS domain containing N termini of the first and second alpha-helices have a number of amino acids critical for the function of the protein; mutations in these regions still allow protein accumulation in the plasma membrane, but the protein is no longer capable of efficiently transporting sulfate into cells. These results suggest that the STAS domain is critical for both the activity and biosynthesis/stability of the transporter, and that STAS sub-domains correlate with these specific functions.

Insights into the acclimation of Chlamydomonas reinhardtii to sulfur deprivation.

During sulfur deprivation, the photosynthetic green alga Chlamydomonas reinhardtii develops a high-affinity sulfate uptake system and increases the expression of genes encoding proteins involved in sulfur assimilation. Although two regulatory elements, SAC1 and SAC3, have been shown to be required for normal acclimation of C. reinhardtii to sulfur deprivation, a number of other regulatory elements appear to also be involved. The molecular mechanisms by which these regulatory elements function are largely unknown. This manuscript presents our current knowledge of sulfur deprivation responses and the regulation of these responses in C. reinhardtii. In addition, we present preliminary results of a sub-saturation screen for novel sulfur acclimation mutants of C. reinhardtii. A speculative model, incorporating the activities of established regulatory elements with putative novel components of the signal transduction pathway(s) is discussed.

Probing the function of STAS domains of the Arabidopsis sulfate transporters.

Sulfate transporters in plants and animals are structurally conserved and have an amino-terminal domain that functions in transport and a carboxyl-terminal region that has been designated the STAS domain. The STAS domain in sulfate transporters has significant similarity to bacterial anti-sigma factor antagonists. To determine if the STAS domain has a role in controlling the activity of sulfate transporters, their stability, or their localization to the plasma membrane, we examined the effect of deleting or modifying the STAS domain of dominant sulfate transporters in roots of Arabidopsis thaliana. The A. thaliana Sultr1;2 and Sultr1;1 sulfate transporters rescue the methionine-dependent growth phenotype of the yeast sulfate transporter mutant strain CP154-7B. Constructs of Sultr1;2 in which the STAS domain was deleted (DeltaSTAS) resulted in synthesis of a truncated polypeptide that was unable to rescue the CP154-7B phenotype. The inability of these constructs to rescue the mutant phenotype probably reflected both low level cellular accumulation of the transporter and the inability of the truncated protein to localize to the plasma membrane. Fusing the STAS domain from other sulfate transporters to Sultr1;2 DeltaSTAS constructs restored elevated accumulation and plasma membrane localization, although the kinetics of sulfate uptake in the transformants were markedly altered with respect to transformants synthesizing wild-type Sultr1;2 protein. These results suggest that the STAS domain is essential, either directly or indirectly, for facilitating localization of the transporters to the plasma membrane, but it also appears to influence the kinetic properties of the catalytic domain of transporters.

Selenate-resistant mutants of Arabidopsis thaliana identify Sultr1;2, a sulfate transporter required for efficient transport of sulfate into roots.

To investigate how plants acquire and assimilate sulfur from their environment, we isolated and characterized two mutants of Arabidopsis thaliana deficient in sulfate transport. The mutants are resistant to selenate, a toxic analogue of sulfate. They are allelic to each other and to the previously isolated sel1 (selenate-resistant) mutants, and have been designated sel1-8 and sel1-9. Root elongation in these mutants is less sensitive to selenate than in wild-type plants. Sulfate uptake into the roots is impaired in the mutants under both sulfur-sufficient and sulfur-deficient conditions, but transport of sulfate to the shoot is not affected. The sel1 mutants contain lesions in the sulfate transporter gene Sultr1;2 located on the lower arm of chromosome 1. The sel1-1, sel1-3 and sel1-8 mutants contain point mutations in the coding sequences of Sultr1;2, while the sel1-9 mutant has a T-DNA insertion in the Sultr1;2 promoter. The Sultr1;2 cDNA derived from wild-type plants is able to complement Saccharomyces cerevisiae mutants defective in sulfate transport, but the Sultr1;2 cDNA from sel1-8 is not. The Sultr1;2 gene is expressed mainly in roots, and accumulation of transcripts increases during sulfate deprivation. Examination of transgenic plants containing the Sultr1;2 promoter fused to the GUS-reporter gene indicates that Sultr1;2 is expressed mainly in the root cortex, the root tip and lateral roots. Weaker expression of the reporter gene was observed in hydathodes, guard cells and auxiliary buds of leaves, and in anthers and the basal parts of flowers. The results indicate that Sultr1;2 is primarily involved in importing sulfate from the environment into the root.