AUF1 cell cycle variations define genomic DNA methylation by regulation of DNMT1 mRNA stability.

DNA methylation is a major determinant of epigenetic inheritance. DNA methyltransferase 1 (DNMT1) is the enzyme responsible for the maintenance of DNA methylation patterns during cell division, and deregulated expression of DNMT1 leads to cellular transformation. We show herein that AU-rich element/poly(U)-binding/degradation factor 1 (AUF1)/heterogeneous nuclear ribonucleoprotein D interacts with an AU-rich conserved element in the 3' untranslated region of the DNMT1 mRNA and targets it for destabilization by the exosome. AUF1 protein levels are regulated by the cell cycle by the proteasome, resulting in cell cycle-specific destabilization of DNMT1 mRNA. AUF1 knock down leads to increased DNMT1 expression and modifications of cell cycle kinetics, increased DNA methyltransferase activity, and genome hypermethylation. Concurrent AUF1 and DNMT1 knock down abolishes this effect, suggesting that the effects of AUF1 knock down on the cell cycle are mediated at least in part by DNMT1. In this study, we demonstrate a link between AUF1, the RNA degradation machinery, and maintenance of the epigenetic integrity of the cell.

Bioprocess development for the production of an antifungal molecule by Bacillus licheniformis BC98.

The optimization of process parameters for the production of an antifungal molecule produced by Bacillus licheniformis BC98 was carried out using novel statistical tools. The parameters studied were pH, temperature and agitation rate. Fed batch cultivations were carried out since the maximum production of the molecule was observed in the late log phase. The statistical design used allows the evaluation of the effects of several different process variables in a single batch. Data from several batches indicated that while the effects of two of the variable factors, viz., temperature and agitation rate, were significant at 95% confidence intervals, the agitation rate was most critical for the production of the molecule, and pH had no significant effect. The cultivation of the bacterium under optimized conditions (fed batch, 150 rpm, 32 degrees C, pH 5.8) resulted in a 30-fold increase compared with that under unoptimized conditions (shake flask, 100 rpm, 29 degrees C, pH 5.8) in the production of the antifungal molecule.

A simple protocol for isolation of fungal DNA.

Genomic DNA was isolated from as little as 2 mg dry biomass of Magnaporthe grisea by microwave treatment within 30 s. The quantity of DNA was good enough for PCR analysis and Dot blot hybridization. This technique can be used for various studies, such as DNA fingerprinting to study the population structure of the phytopathogen in different regions, and for a quick screening of M. grisea transformants.