GENETIC VARIATION IN THE TEMPERATURE DEPENDENCE OF LIVER MICROSOMAL CYP2E1 ACTIVITY, WITHIN AND BETWEEN SPECIES OF THE VIVIPAROUS FISH POECILIOPSIS

摘要

The temperature dependence of liver microsomal CYP2E1 (cytochrome P450 2E1) activity was examined in selected genotypes of the viviparous fish Poeciliopsis. Activity of this enzyme, as a function of incubation temperature, was determined by measuring 6-OH-chlorzoxazone formation from chlorzoxazone, a specific CYP2E1 substrate. Chlorzoxazone-6-hydroxylase activity was examined among five species of Poeciliopsis, as well as among nine genotypes within a species, P. monacha. Among Poeciliopsis genotypes, P. monacha contained the greatest activity, 9.5 ± 1.5 U (where 1 U is defined as 1 μg 6-OH-chlorzoxazone produced/min/mg protein; n = 6) with a temperature optimum (T_o) of 25℃. The lowest activity was in P. occidentalis, 0.65 ± 0.11 U, with a T_o of 27 to 28℃; P. prolifica, P. fasciata, P. lucida, and P. viriosa had intermediate levels of activities, 1.1 to 5.5 U, and T_o from 25 to 31℃. To determine if metabolic differences exist within species, enzyme activity was examined from nine genotypes of P. monacha by comparing expression among P. monacha-lucida hybrids. These hybrids were given identical paternal genomes of lucida but retained the original maternal monacha genomes sampled from wild gene pools. The greatest activity was found in genotype T70-3 P. Cw, 3.6 ± 0.1 U, at a T_o of 29℃, and the lowest was in genotype SV73-7s, 0.40 ±0.12 U, at a T_o of 27℃. The other naturally occurring genotypes, M65-24, M65-26, SV73-7v, as well as the laboratory-produced synthetic hybrids, Syn4 and Syn5, had intermediate activities, 0.73 ± 0.38 to 2.1 ± 0.69 U, and T_o of 25 to 27℃. No hybrid had activity levels as high as the maternal parent, P. monacha, and only one had a T_o as low as either parent. Apparently the genes involved in xenobiotic activity vary widely among the closely related species of Poeciliopsis but also within species, suggesting that these phenotypes can be acted upon by natural selection.