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DIFFERENCES IN THE STRUCTURE AND FUNCTION OF FATHEAD MINNOW AND HUMAN ERA: IMPLICATIONS FOR IN VITRO TESTING OF ENDOCRINE DISRUPTING CHEMICALS
Citation:
RIDER, C. V., P. C. HARTIG, M. C. CARDON, AND V. S. WILSON. DIFFERENCES IN THE STRUCTURE AND FUNCTION OF FATHEAD MINNOW AND HUMAN ERA: IMPLICATIONS FOR IN VITRO TESTING OF ENDOCRINE DISRUPTING CHEMICALS. Presented at SOT Annual Meeting, Seattle, WA, March 16 - 20, 2008.
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Mammalian receptors and assay systems are generally used for in vitro analysis of endocrine disrupting chemicals (EDC) with the assumption that minor differences in amino acid sequences among species do not translate into significant differences in receptor function. We have found that the fathead minnow estrogen receptor alpha (fER) differs from the human estrogen receptor alpha (hER) in terms of both the amino acid sequence of the ligand binding pocket and the affinity of some EDCs for the receptor. The fER was previously isolated from a cDNA library and sequenced. Chemicals are currently being tested for their affinity to either fER or hER in two binding assays to compare results from whole cell versus cell-free binding systems. The fER had reduced ligand binding at a higher temperature (37°C), whereas, ligand affinity for the hER was similar at both 37°C and room temperature. A few chemicals displayed notable differences in binding affinities among fER and hER. These included the antiestrogen ICI, dibutyl phthalate (DBP), and benzyl butyl phthalate (BBP) (fER IC50s = 41.6±6.7nM, 18.1±7.3M, and 8.2±2.1M, respectively; hER IC50s = 192.6±23.1nM for ICI and could not be calculated for DPB and BBP due to incomplete binding curves). Confirmatory studies in the cell-free binding system are in progress. If confirmed, these results suggest that studies using mammalian receptors and cell systems for in vitro screening and testing of chemicals may not always reflect EDC affinity for receptors among lower vertebrates.