The <i>Trypanosoma cruzi</i> (<i>T. cruzi</i>) parasite is the cause of Chagas disease, a neglected disease endemic in South America. The life cycle of the <i>T. cruzi</i> parasite is complex and includes transitions between distinct life stages. This change in phenotype (without a change in genotype) could be controlled by epigenetic regulation, and might involve the bromodomain-containing factors 1-5 (<i>Tc</i>BDF1-5). However, little is known about the function of the <i>Tc</i>BDF1-5. Here we describe a fragment-based approach to identify ligands for <i>T. cruzi</i> bromodomain-containing factor 3 (<i>Tc</i>BDF3). We expressed a soluble construct of <i>Tc</i>BDF3 in <i>E. coli</i>, and used this to develop a range of biophysical assays for this protein. Fragment screening identified twelve compounds that bind to the <i>Tc</i>BDF3 bromodomain. Based on this screen, we developed functional ligands containing a fluorescence or <sup>19</sup>F reporter group, and a photo-crosslinking probe for <i>Tc</i>BDF3. These tools compounds will be invaluable in future studies on the function of <i>Tc</i>BDF3 and will provide insight into the biology of <i>T. cruzi</i>.
Sodium fluoroacetate (1080) is a highly toxic metabolic poison that has the potential because of its lack of defined color, odor, and taste and its high water solubility to be intentionally or unintentionally ingested through food adulteration. Although the mechanism of action for 1080 has been known since the 1950s, no known antidote exists. In an effort to better understand the cardiopulmonary impacts of 1080, we utilized whole-body plethysmography and telemeterized Sprague-Dawley rats which allowed for the real-time measurement of respiratory and cardiac parameters following exposure using a non-invasive assisted-drinking method. Overall, the animals showed marked depression of respiratory parameters over the course of 24 h post-exposure and the development of hemorrhage in the lung tissue. Tidal volume was reduced by 30% in males and 60% in females at 24 h post-exposure, and respiratory frequency was significantly depressed as well. In telemeterized female rats, we observed severe cardiac abnormalities, highlighted by a 50% reduction in heart rate, 75% reduction in systolic blood pressure, and a 3.5-fold lengthening of the QRS interval over the course of 24 h. We also observed a reduction in core body temperature of nearly 15°C. Our study was able to describe the severe and pronounced effects of sodium fluoroacetate poisoning on cardiopulmonary function, the results of which indicate that both tissue specific and systemic deficits contribute to the toxicological progression of 1080 intoxication ABOUT THE AUTHOR Bryan J. McCranor is a research biochemist at the US Army Medical Research Institute of Chemical Defense where he is a principal investigator with the Inhalation Toxicology Team. Overall, his research focuses on the development of novel prophylactic, therapeutic and medical countermeasure treatments for exposure to chemical threat agents and toxic industrial compounds. His work centers on the identification of toxicological characteristics of toxic chemicals, investigation of potential novel therapeutic targets, and advancement of medical countermeasures.
The <i>Trypanosoma cruzi</i> (<i>T. cruzi</i>) parasite is the cause of Chagas disease, a neglected disease endemic in South America. The life cycle of the <i>T. cruzi</i> parasite is complex and includes transitions between distinct life stages. This change in phenotype (without a change in genotype) could be controlled by epigenetic regulation, and might involve the bromodomain-containing factors 1-5 (<i>Tc</i>BDF1-5). However, little is known about the function of the <i>Tc</i>BDF1-5. Here we describe a fragment-based approach to identify ligands for <i>T. cruzi</i> bromodomain-containing factor 3 (<i>Tc</i>BDF3). We expressed a soluble construct of <i>Tc</i>BDF3 in <i>E. coli</i>, and used this to develop a range of biophysical assays for this protein. Fragment screening identified twelve compounds that bind to the <i>Tc</i>BDF3 bromodomain. Based on this screen, we developed functional ligands containing a fluorescence or <sup>19</sup>F reporter group, and a photo-crosslinking probe for <i>Tc</i>BDF3. These tools compounds will be invaluable in future studies on the function of <i>Tc</i>BDF3 and will provide insight into the biology of <i>T. cruzi</i>.
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