Adverse effects of bisphenol A (BPA) on the dopamine system in two distinct cell models and corpus striatum of the Sprague-Dawley rat.

The aim of this study was to examine the effects of bisphenol A (BPA) on the brain dopamine (DA) system utilizing both in vitro models (GH3 cells, a rat pituitary cell line, and SH-SY5Y cells, a human neuroblastoma cell line) and an animal model such as Sprague-Dawley (SD) rats. First, cellular DA uptake was measured 2 or 8 h following BPA exposure (0.1-400 µM) in SH-SY5Y cells, where a significant increase in DA uptake was noted. BPA exerted no marked effect on dopamine active transporter levels in GH3 cells exposed for 8 or 24 h. However, SH-SY5Y cells displayed an increase in dopamine transporter (DAT) levels following 24 h of exposure to BPA. In contrast to DAT levels, BPA exposure produced no marked effect on DA D1 receptor levels in SH-SY5Y cells, yet a significant decrease in GH3 cells following both 8- and 24-h exposure periods was noted, suggesting that BPA exerts differential effects dependent upon cell type. BPA produced no significant effects on prolactin levels at 2 h, but a marked fall occurred at 24 h of exposure in GH3 cells. Finally, to examine the influence of dietary developmental exposure to BPA on brain DA levels in F1 offspring, SD rats were exposed to BPA (0.5-20 mg/kg) through maternal transfer and/or diet and striatal DA levels were measured on postnatal day (PND) 60 using high-performance liquid chromatography (HPLC). Data demonstrated that chronic exposure to BPA did not significantly alter striatal DA levels in the SD rat.

Critical Components of the Conjugation Machinery of the Integrative and Conjugative Element ICEBs1 of Bacillus subtilis.

UNLABELLED: Conjugation, or mating, plays a profound role in bacterial evolution by spreading genes that allow bacteria to adapt to and colonize new niches. ICEBs1, an integrative and conjugative element of Bacillus subtilis, can transfer itself and mobilize resident plasmids. DNA transfer is mediated by a type IV secretion system (T4SS). Characterized components of the ICEBs1 T4SS include the conserved VirB4-like ATPase ConE, the bifunctional cell wall hydrolase CwlT, and the presumed VirD4-like coupling protein ConQ. A fusion of ConE to green fluorescent protein (GFP) localizes to the membrane preferentially at the cell poles. One or more ICEBs1 proteins are required for ConE's localization at the membrane, as ConE lacks predicted transmembrane segments and ConE-GFP is found dispersed throughout the cytoplasm in cells lacking ICEBs1. Here, we analyzed five ICEBs1 genes to determine if they are required for DNA transfer and/or ConE-GFP localization. We found that conB, conC, conD, and conG, but not yddF, are required for both ICEBs1 transfer and plasmid mobilization. All four required genes encode predicted integral membrane proteins. conB and, to some extent, conD were required for localization of ConE-GFP to the membrane. Using an adenylate cyclase-based bacterial two-hybrid system, we found that ConE interacts with ConB. We propose a model in which the ICEBs1 conjugation machinery is composed of ConB, ConC, ConD, ConE, ConG, CwlT, ConQ, and possibly other ICEBs1 proteins, and that ConB interacts with ConE, helping to recruit and/or maintain ConE at the membrane. IMPORTANCE: Conjugation is a major form of horizontal gene transfer and has played a profound role in bacterial evolution by moving genes, including those involved in antibiotic resistance, metabolism, symbiosis, and infectious disease. During conjugation, DNA is transferred from cell to cell through the conjugation machinery, a type of secretion system. Relatively little is known about the conjugation machinery of Gram-positive bacteria. Here, we analyzed five genes of the integrative and conjugative element ICEBs1 of Bacillus subtilis. Our research identifies four new components of the ICEBs1 conjugation machinery (ConB, ConC, ConD, and ConG) and shows an interaction between ConB and ConE that is required for ConE to associate with the cell membrane.

ATP sequestration by a synthetic ATP-binding protein leads to novel phenotypic changes in Escherichia coli.

Artificial proteins that bind key metabolites with high affinity and specificity hold great promise as new tools in synthetic biology, but little has been done to create such molecules and examine their effects on living cells. Experiments of this kind have the potential to expand our understanding of cellular systems, as certain phenotypes may be physically realistic but not yet observed in nature. Here, we examine the physiology and morphology of a population of Escherichia coli as they respond to a genetically encoded, non-biological ATP-binding protein. Unlike natural ATP-dependent proteins, which transiently bind ATP during metabolic transformations, the synthetic protein DX depletes the concentration of intracellular ATP and ADP by a mechanism of protein-mediated ligand sequestration. The resulting ATP/ADP imbalance leads to an adaptive response in which a large population of bacilli cells transition to a filamentous state with dense lipid structures that segregate the cells into compartmentalized units. A wide range of biochemical and microscopy techniques extensively characterized these novel lipid structures, which we have termed endoliposomes. We show that endoliposomes adopt well-defined box-like structures that span the full width of the cell but exclude the synthetic protein DX. We further show that prolonged DX exposure causes a large fraction of the population to enter a viable-but-non-culturable state that is not easily reversed. Both phenotypes correlate with strong intracellular changes in ATP and ADP concentration. We suggest that artificial proteins, such as DX, could be used to control and regulate specific targets in metabolic pathways.