BES1/BZR1 Homolog 3 cooperates with E3 ligase AtRZF1 to regulate osmotic stress and brassinosteroid responses in Arabidopsis.

Proline (Pro) metabolism plays important roles in protein synthesis, redox balance, and abiotic stress response. However, it is not known if cross-talk occurs between proline and brassinosteroid (BR) signaling pathways. Here, an Arabidopsis intergenic enhancer double mutant, namely proline content alterative 41 (pca41), was generated by inserting a T-DNA tag in the Arabidopsis thaliana ring zinc finger 1 (atrzf1 ) mutant background. pca41 had a T-DNA inserted at the site of the gene encoding BES1/BZR1 Homolog 3 (BEH3). pca41 has a drought-insensitive phenotype that is stronger than atrzf1 under osmotic stress, including high Pro accumulation and decreased amounts of reactive oxygen species. Analysis of physiological, genetic, and molecular networks revealed that negative regulation of BEH3 during abiotic stress was linked to the BR signaling pathway. Our data also suggest that AtRZF1, an E3 ubiquitin ligase, might control osmotic stress, abscisic acid, and BR responses in a BEH3-dependent manner. Under darkness, pca41 displays a long hypocotyl phenotype, which is similar to atrzf1 and beh3, suggesting that BEH3 acts in the same pathway as AtRZF1. Overexpression of BEH3 results in an osmotic stress-sensitive phenotype, which is reversed by exogenous BR application. Taken together, our results indicate that AtRZF1 and BEH3 may play important roles in the osmotic stress response via ubiquitination and BR signaling.

Loss of Arabidopsis Halotolerance 2-like (AHL), a 3'-phosphoadenosine-5'-phosphate phosphatase, suppresses insensitive response of Arabidopsis thaliana ring zinc finger 1 (atrzf1) mutant to abiotic stress.

KEY MESSAGE: Destruction of PAP phosphatase AHL suppresses atrzf1 phenotype in abiotic stress responses. AHL plays an intermediate role in the regulation of proline accumulation by PAP nucleotidase. Proline (Pro) metabolism is important for environmental responses, plant development, and growth. However, the role of Pro in abiotic stress process is unclear. Using atrzf1 (Arabidopsis thaliana ring zinc finger 1) mutant as a parental line for T-DNA tagging mutagenesis, we identified a suppressor mutant designated as proline content alterative 17 (pca17) that suppressed insensitivity of atrzf1 to abiotic stresses during early seedling growth. Pro content of pca17 was lower than that in both wild type (WT) and atrzf1 while complementary lines were less sensitive to abscisic acid (ABA) and abiotic stresses compared to WT. Thermal Asymmetric Interlaced (TAIL)-PCR of pca17 showed that T-DNA was inserted at site of At5g54390 (AHL for Arabidopsis Halotolerance 2-like) encoding 3'-phosphoadenosine-5'-phosphate (PAP) phosphatase. Under drought stress condition, products of sulfate metabolism such as PAP and adenosine monophosphate were significantly lower in pca17 than those in WT and atrzf1. Furthermore, pca17 showed significantly higher levels of several important drought parameters including malondialdehyde, ion leakage, and water loss than WT and atrzf1. Fluorescence signal of green fluorescent protein (GFP)-tagged AHL was quite strong in nuclei of the root and guard cells of transgenic seedlings. Additionally, AHL promoter-ß-glucuronidase (GUS) construct revealed substantial gene expression in vasculature tissues and pollen. Collectively, these findings demonstrate that pca17 acts as a dominant suppressor mutant of atrzf1 in abiotic stress response by modulating proline and sulfate metabolism.

Isolation and characterization of a thermophilic chain elongating bacterium that produces the high commodity chemical n-caproate from polymeric carbohydrates.

A thermophilic chain elongating bacterium, strain MDTJ8, was isolated from a thermophilic acidogenic anaerobic digestor producing n-caproate from human waste, growing optimally at 50-55 °C and pH 6.5. 16S rRNA gene analysis suggests that MDTJ8 represents a new species/genus within a group currently composed of mesophilic chain elongators of the Oscillospiraceae family. Genome analysis showed that strain MDTJ8 contains homologues of genes encoding for chain elongation and energy conservation but also indicated n-caproate production from carbohydrates including polymeric substances. This was confirmed by culturing experiments in which MDTJ8 converted, at pH 6.5 and 55 °C, mono-, di- and polymeric carbohydrates (starch and hemicellulose) to n-caproate reaching concentrations up to 283 mg/L and accounting for up to 10 % of the measured fermentation products. MDTJ8 is the first axenic organism that thermophilically performs chain elongation, opening doors to understand and intensify thermophilic bioprocesses targeting anaerobic digestion towards the production of the value-added chemical n-caproate.

proline content alterative 17 (pca17) is involved in glucose response through sulfate metabolism-mediated pathway.

Sulfate metabolism and glucose (Glc) signaling are important processes required for plant growth, development, and environmental responses. However, whether sulfate metabolism is involved in Arabidopsis response to Glc stress remains largely unclear. Recently, we have found that proline content alterative 17 (pca17) is a double-mutant line in which both AtRZF1 (for Arabidopsis thaliana Ring Zinc Finger 1) and AHL (for Arabidopsis Halotolerance 2-like) genes are mutated. It was found that insensitive response of atrzf1 mutant to abiotic stresses was suppressed in pca17 mutant by regulating proline metabolism. Here, pca17 appeared to have sensitive response to Glc treatment by reducing cysteine (Cys) and adenosine monophosphate (AMP) contents in sulfate metabolism. Under Glc treatment, transcript levels of sulfate metabolism-related genes were significantly lower in pca17 than those in wild-type (WT) and atrzf1. Furthermore, AHL-overexpressing transgenic lines displayed more insensitive phenotypes than WT during Glc condition while ahl RNAi lines exhibited sensitive responses based on several parameters, including seed germination rate, cotyledon greening percentage, root elongation, and fresh weight. Interestingly, the pca17 phenotype in applied AMP with Glc treatment was similar to atrzf1 phenotype. Taken together, our results indicate that AHL is involved in Glc response by modulating sulfate metabolism in Arabidopsis.