ABSTRACT
Understanding how drugs work in vivo is critical for drug design and for maximizing the potential of currently available drugs. 5-nitrofurans are a class of prodrugs widely used to treat bacterial and trypanosome infections, but despite relative specificity, 5-nitrofurans often cause serious toxic side effects in people. Here, we use yeast and zebrafish, as well as human in vitro systems, to assess the biological activity of 5-nitrofurans, and we identify a conserved interaction between aldehyde dehydrogenase (ALDH) 2 and 5-nitrofurans across these species. In addition, we show that the activity of nifurtimox, a 5-nitrofuran anti-trypanosome prodrug, is dependent on zebrafish Aldh2 and is a substrate for human ALDH2. This study reveals a conserved and biologically relevant ALDH2-5-nitrofuran interaction that may have important implications for managing the toxicity of 5-nitrofuran treatment.
Subject(s)
Aldehyde Dehydrogenase/metabolism , Melanocytes/drug effects , Nitrofurans/pharmacology , Saccharomyces cerevisiae/drug effects , Aldehyde Dehydrogenase, Mitochondrial , Animals , Dose-Response Relationship, Drug , Humans , Models, Molecular , Molecular Structure , Nitrofurans/chemistry , Recombinant Proteins/metabolism , Saccharomyces cerevisiae/metabolism , Species Specificity , Structure-Activity Relationship , Zebrafish/embryologyABSTRACT
Small molecules complement genetic mutants and can be used to probe pigment cell biology by inhibiting specific proteins or pathways. Here, we present the results of a screen of active compounds for those that affect the processes of melanocyte and iridophore development in zebrafish and investigate the effects of a few of these compounds in further detail. We identified and confirmed 57 compounds that altered pigment cell patterning, number, survival, or differentiation. Additional tissue targets and toxicity of small molecules are also discussed. Given that the majority of cell types, including pigment cells, are conserved between zebrafish and other vertebrates, we present these chemicals as molecular tools to study developmental processes of pigment cells in living animals and emphasize the value of zebrafish as an in vivo system for testing the on- and off-target activities of clinically active drugs.
Subject(s)
Metabolic Networks and Pathways/drug effects , Pigmentation/drug effects , Small Molecule Libraries/analysis , Small Molecule Libraries/pharmacology , Zebrafish/metabolism , Animals , Cell Count , Chromatophores/cytology , Chromatophores/drug effects , Cyclooxygenase Inhibitors/pharmacology , Drug Evaluation, Preclinical , Embryo, Nonmammalian/drug effects , Embryo, Nonmammalian/metabolism , Heterocyclic Compounds, 3-Ring/pharmacology , Melanocytes/cytology , Melanocytes/drug effects , Phenotype , Purines/pharmacology , Pyrimidinones/pharmacology , Roscovitine , Tyrphostins/pharmacology , Zebrafish/embryologyABSTRACT
Coordination of cell proliferation and differentiation is crucial for tissue formation, repair and regeneration. Some tissues, such as skin and blood, depend on differentiation of a pluripotent stem cell population, whereas others depend on the division of differentiated cells. In development and in the hair follicle, pigmented melanocytes are derived from undifferentiated precursor cells or stem cells. However, differentiated melanocytes may also have proliferative capacity in animals, and the potential for differentiated melanocyte cell division in development and regeneration remains largely unexplored. Here, we use time-lapse imaging of the developing zebrafish to show that while most melanocytes arise from undifferentiated precursor cells, an unexpected subpopulation of differentiated melanocytes arises by cell division. Depletion of the overall melanocyte population triggers a regeneration phase in which differentiated melanocyte division is significantly enhanced, particularly in young differentiated melanocytes. Additionally, we find reduced levels of Mitf activity using an mitfa temperature-sensitive line results in a dramatic increase in differentiated melanocyte cell division. This supports models that in addition to promoting differentiation, Mitf also promotes withdrawal from the cell cycle. We suggest differentiated cell division is relevant to melanoma progression because the human melanoma mutation MITF(4T)(Δ)(2B) promotes increased and serial differentiated melanocyte division in zebrafish. These results reveal a novel pathway of differentiated melanocyte division in vivo, and that Mitf activity is essential for maintaining cell cycle arrest in differentiated melanocytes.
Subject(s)
Cell Division , Melanocytes/cytology , Melanocytes/metabolism , Microphthalmia-Associated Transcription Factor/metabolism , Mutation , Zebrafish Proteins/metabolism , Zebrafish/metabolism , Animals , Cell Differentiation , Gene Expression Regulation, Developmental , Humans , Microphthalmia-Associated Transcription Factor/genetics , Zebrafish/embryology , Zebrafish/genetics , Zebrafish Proteins/geneticsABSTRACT
In the past two decades, zebrafish genetic screens have identified a wealth of mutations that have been essential to the understanding of development and disease biology. More recently, chemical screens in zebrafish have identified small molecules that can modulate specific developmental and behavioural processes. Zebrafish are a unique vertebrate system in which to study chemical genetic systems, identify drug leads, and explore new applications for known drugs. Here, we discuss some of the advantages of using zebrafish in chemical biology, and describe some important and creative examples of small molecule screening, drug discovery and target identification.
