Crystal structure of the N-terminal domain of the yeast general corepressor Tup1p and its functional implications.

The yeast Cyc8p-Tup1p protein complex is a general transcriptional corepressor of genes involved in many different physiological processes. Herein, we present the crystal structure of the Tup1p N-terminal domain (residues 1-92), essential for Tup1p self-assembly and interaction with Cyc8p. This domain tetramerizes to form a novel antiparallel four-helix bundle. Coiled coil interactions near the helical ends hold each dimer together, whereas interdimeric association involves only two sets of two residues located toward the chain centers. A mutagenesis study confirmed that the nonpolar residues responsible for the association of the protomers as dimers are also required for transcriptional repression. An additional structural study demonstrated that the domain containing an Leu(62) → Arg mutation that had been shown not to bind Cyc8p exhibits an altered structure, distinct from the wild type. This altered structure explains why the mutant cannot bind Cyc8p. The data presented herein highlight the importance of the architecture of the Tup1p N-terminal domain for self-association.

Interspecies interactions are an integral determinant of microbial community dynamics.

This study investigated the factors that determine the dynamics of bacterial communities in a complex system using multidisciplinary methods. Since natural and engineered microbial ecosystems are too complex to study, six types of synthetic microbial ecosystems (SMEs) were constructed under chemostat conditions with phenol as the sole carbon and energy source. Two to four phenol-degrading, phylogenetically and physiologically different bacterial strains were used in each SME. Phylogeny was based on the nucleotide sequence of 16S rRNA genes, while physiologic traits were based on kinetic and growth parameters on phenol. Two indices, J parameter and "interspecies interaction," were compared to predict which strain would become dominant in an SME. The J parameter was calculated from kinetic and growth parameters. On the other hand, "interspecies interaction," a new index proposed in this study, was evaluated by measuring the specific growth activity, which was determined on the basis of relative growth of a strain with or without the supernatant prepared from other bacterial cultures. Population densities of strains used in SMEs were enumerated by real-time quantitative PCR (qPCR) targeting the gene encoding the large subunit of phenol hydroxylase and were compared to predictions made from J parameter and interspecies interaction calculations. In 4 of 6 SEMs tested the final dominant strain shown by real-time qPCR analyses coincided with the strain predicted by both the J parameter and the interspecies interaction. However, in SMEII-2 and SMEII-3 the final dominant Variovorax strains coincided with prediction of the interspecies interaction but not the J parameter. These results demonstrate that the effects of interspecies interactions within microbial communities contribute to determining the dynamics of the microbial ecosystem.