Characterization of Arabidopsis AtUGT85A and AtGUS gene families and their expression in rapidly dividing tissues.

In humans, uridine 5'-diphosphate glucuronosyltransferase (UGT) operates in opposition to glucuronidase (GUS) to control activity of diverse metabolites such as hormones by reversible conjugation with glucuronic acid. Previous data revealed that, as in mammals, these enzymes are required for plant life in that a UGT from Pisum sativum (PsUGT1) controls plant development by opposing endogenous GUS activity thereby modulating the duration of the cell cycle. Here we report that a small family of genes (AtUGT85A1, 2, 3, 4, 5, and 7) homologous to pea PsUGT1 exists in the Arabidopsis genome. The AtUGT85A-encoded proteins are predicted to be membrane-associated enzymes. Three genes (AtGUS1, AtGUS2, and AtGUS3) that are homologous to a GUS-encoding gene from Scutellaria baicalensis were identified. The AtGUS-encoded proteins are predicted to be secretory (AtGUS1) and membrane-associated (AtGUS2 and AtGUS3) enzymes. Both AtUGT85A and AtGUS genes, like PsUGT1, exhibit localized, tissue-specific expression, mainly in areas of active cell division with possible involvement in cell cycle regulation.

Characterization, cloning and expression of the ferritin gene from the Korean polychaete, Periserrula leucophryna.

Ferritin is a major eukaryotic protein and in humans is the protein of iron storage. A partial gene fragment of ferritin (255 bp) taken from the total RNA of Periserrula leucophryna, was amplified by RT-PCR using oligonucleotide primers designed from the conserved metal binding domain of eukaryotic ferritin and confirmed by DNA sequencing. Using the 32P-labeled partial ferritin cDNA fragment, 28 different clones were obtained by the screening of the P. leucophryna cDNA library prepared in the Uni-ZAP XR vector, sequenced and characterized. The longest clone was named the PLF (Periserrula leucophryna ferritin) gene and the nucleotide and amino acid sequences of this novel gene were deposited in the GenBank databases with accession numbers DQ207752 and ABA55730, respectively. The entire cDNA of PLF clone was 1109 bp (CDS: 129-653), including a coding nucleotide sequence of 525 bp, a 5'-untranslated region of 128 bp, and a 3'-noncoding region of 456 bp. The 5'-UTR contains a putative iron responsive element (IRE) sequence. Ferritin has an open reading frame encoding a polypeptide of 174 amino acids including a hydrophobic signal peptide of 17 amino acids. The predicted molecular weights of the immature and mature ferritin were calculated to be 20.3 kDa and 18.2 kDa, respectively. The region encoding the mature ferritin was subcloned into the pT7-7 expression vector after PCR amplification using the designed primers and included the initiation and termination codons; the recombinant clones were expressed in E. coli BL21(DE3) or E. coli BL21(DE3)pLysE. SDS-PAGE and western blot analysis showed that a ferritin of approximately 18 kDa (mature form) was produced and that by iron staining in native PAGE, it is likely that the recombinant ferritin is correctly folded and assembled into a homopolymer composed of a single subunit.

Flavonoids: from cell cycle regulation to biotechnology.

Flavonoids have been proposed to play diverse roles in plant growth and development, including defense, symbiosis, pollen development and male fertility, polar auxin transport, and protection against ultraviolet radiation. Recently, a new role in cell cycle regulation has emerged. Genetic alteration of glucuronide metabolism by altered expression of a Pisum sativum UDP-glucuronosyltransferase (PsUGT1) results in an altered cell cycle in pea, alfalfa, and Arabidopsis. In alfalfa, altered expression of PsUGT1 results in accumulation of a flavonoid-like compound that suppresses growth of cultured cells. The results are consistent with the hypothesis that PsUGT1 functions by controlling cellular levels of a factor controlling cell cycle (FCC).

Removal of organic matter and nitrogen from river water in a model floodplain.

A significant improvement in river water quality cannot be expected unless nonpoint-source contaminants are treated in addition to the further treatment of point-source contaminants. If river water is sprayed over a floodplain, the consequent water filtration through the sediment profile can simultaneously remove organic matter and nitrogen in the water through aerobic and denitrifying reactions. This hypothesis was tested using lysimeters constructed from polyvinyl chloride (PVC) pipe (150 cm long, 15 cm in diameter) packed with loamy sand floodplain sediment. Water was applied to the top of the lysimeters at three different flow rates (48, 54, and 68 mm d(-1)). Concentrations of NO3 and dissolved oxygen (DO), chemical oxygen demand (COD), and redox potential (Eh) in the water were measured as functions of depth after the system reached steady states for both water flow and reactions. At the rate of 68.0 mm d(-1), a reducing condition for denitrification developed below the 5-cm depth due to the depletion of O2 by organic matter degradation in the surface oxidizing layer; Eh and DO were below 205 mV and 0.4 mg L(-1), respectively. At a depth of 70 cm, COD and NO3-N concentration decreased to 5.2 and 3.8 mg L(-1) from the respective influent concentrations of 17.1 and 6.2 mg L(-1). Most biodegradable organic matter was removed during flow and further removal of NO3 was limited by the lack of an electron donor (i.e., organic matter). These results indicate that the floodplain filtration technique has great promise for treatment of contaminated river water.

Electron affinity coefficients of nitrogen oxides and biodegradation kinetics in denitrification of contaminated stream water.

During the dry season in Korea, rivers become more vulnerable to contamination by biochemical oxygen demand (BOD) and nitrogen. It is hypothesized that the natural characteristics of the streams in Korea allow the contaminated water to be treated at the tributaries. Down-stream river water quality in Korea may be improved by spraying the contaminated stream water from the tributaries over the surrounding floodplains. The consequent water filtration through the soil could remove the contaminants through aerobic and denitrifying reactions. In this study, the kinetics parameters of the denitrifying reaction in floodplain filtration were determined using contaminated stream water. For the electron donor the Monod kinetics was used, while the competitive Michaelis-Menten model was employed for the electron acceptors. The parameters to the competitive Michaelis-Menten model were found using continuous denitrifying reactions, instead of the batch reactions employed in previous studies, to match the conditions needed to apply the competitive Michaelis-Menten kinetics. From the result, it was found that continuous reactions as well as batch reactions could be used to determine the affinity coefficients in denitrification. The results of this study also showed that the affinity coefficient of NO2, using continuous reactions, was similar to that of other studies in the literature found via batch reactions, whereas the affinity coefficient of N2O was much larger than that acquired with batch reactions. The parameters obtained in this study will be used in future work to simulate the contaminant behaviors during floodplain filtration using a mathematical model.