Cryptic speciation in the field vole: a multilocus approach confirms three highly divergent lineages in Eurasia

Species are generally described from morphological features, but there is growing recognition of sister forms that show substantial genetic differentiation without obvious morphological variation and may therefore be considered ‘cryptic species’. Here, we investigate the field vole (Microtus agrestis), a Eurasian mammal with little apparent morphological differentiation but which, on the basis of previous sex‐linked nuclear and mitochondrial DNA (mtDNA) analyses, is subdivided into a Northern and a Southern lineage, sufficiently divergent that they may represent two cryptic species. These earlier studies also provided limited evidence for two major mtDNA lineages within Iberia. In our present study, we extend these findings through a multilocus approach. We sampled 163 individuals from 46 localities, mainly in Iberia, and sequenced seven loci, maternally, paternally and biparentally inherited. Our results show that the mtDNA lineage identified in Portugal is indeed a distinct third lineage on the basis of other markers as well. In fact, multilocus coalescent‐based methods clearly support three separate evolutionary units that may represent cryptic species: Northern, Southern and Portuguese. Divergence among these units was inferred to have occurred during the last glacial period; the Portuguese lineage split occurred first (estimated at c. 70 000 bp ), and the Northern and Southern lineages separated at around the last glacial maximum (estimated at c. 18 500 bp ). Such recent formation of evolutionary units that might be considered species has repercussions in terms of understanding evolutionary processes and the diversity of small mammals in a European context.     

Fast product formation and slow product release are important features in a hysteretic reaction mechanism of glutathione transferase T2-2.

The reaction mechanism of rat glutathione transferase T2-2 has been studied using pre-steady-state and steady-state kinetics. Several parts of the catalytic cycle including binding of substrates, product formation, and product release were investigated. Under saturating conditions, a two-step product release was found to be rate limiting in the enzyme-catalyzed reactions between the nucleophilic substrate glutathione and either of the two electrophilic substrates 1-menaphthyl sulfate and 4-nitrobenzyl chloride. The rate constant for pre-steady-state product formation on rat glutathione transferase T2-2 has an observed pK(a) value of 5.7 apparently due to ionization of the sulfhydryl group of glutathione. This rate constant is approximately 2 orders of magnitude higher than k(cat) at pH values of >6. It can be predicted from the pH dependence that product formation would be the sole rate-limiting step at pH values of <3. A hysteretic mechanism of rGST T2-2 is proposed based on a slow conformational transition detected in pre-steady-state displacement experiments.     

Ethological study of early language

Sign language studies of cross-fostered chimpanzees measure the effect of special rearing conditions on the development of very young chimpanzees. Cross-fostered chimpanzees, like human children, develop gradually in a process that takes many years. Here we discuss details of the procedure, the overlap between human and chimpanzee infants in the contents of the first 50-item vocabularies, and the ways in which the signs of the chimpanzees exhibit the fuzziness of natural language categories. We also compare the cross-fostering approach with more traditional modular approaches to the study of language-like behavior in nonhuman animals. Project Washoe was originally supported by grants MH-12154 from the National Institute of Mental Health and GB-7432 from the National Science Foundation. We gratefully acknowledge this support and the support that later sign language studies of chimpanzees have received since then from NIH, NSF, the National Geographic Society, the Grant Foundation, the Spencer Foundation, the University of Nevada, and the UNR Foundation.     

Mutagenicity of methylisocyanate and its reaction products to cultured mammalian cells

Methylisocyanate (MIC) induced mutagenic responses in the absence of exogenous activation in the mouse lymphoma cell forward mutation assay at concentrations as low as 8-24 microM. MIC produced predominantly small mutant colonies, suggesting the possibility of clastogenic activity. The intermediate hydrolysis product, methylamine, was also mutagenic without exogenous activation but required several hundred-fold higher concentrations (ca. 3 mM). N,N'-Dimethylurea, the final product in the reaction of methylisocyanate and water, was totally refractory in either the presence or absence of S9 for concentrations up to 57 mM (5 mg/ml). The ethyl ester of N-methylcarbamic acid was also tested since it was the only available analogue to the highly reactive N-methylcarbamic acid intermediate. This compound was mutagenic only in the presence of S9 at doses exceeding 5-40 microM, which suggested the possibility that the free acid, produced by enzymatic hydrolysis, is also mutagenic. The mutagenic activity of the ester resulted solely in the production of small mutant colonies.     

The effect of phenformin and other adenosine triphosphate (ATP)-lowering agents on insulin binding to IM-9 human cultured lymphocytes

In the present study, we investigated the mechanism by which the antidiabetic drug phenformin increases insulin binding to its receptors in IM-9 human cultured lymphocytes. After a 24-hr preincubation, phenformin induced a twofold increase in specific 125I-insulin binding, and removal of phenformin was followed 6 hr later by a return in binding to control levels. This effect of phenformin on insulin binding was not a consequence of either inhibition of cell growth, changes in cellular cyclic adenosine monophosphate (AMP) levels, or changes in guanosine triphosphate (GTP) content. Since phenformin is known to inhibit various aspects of cellular energy metabolism, the relationship between 125I-insulin binding and energy metabolism in IM-9 cells was investigated. The phenformin-induced increase in insulin binding to IM-9 cells was related to a time- and dose-dependent decrease in ATP levels. Other agents that lowered ATP levels, including antimycin, dinitrophenol, and 2-deoxyglucose, also raised insulin binding. These studies indicated, therefore, that phenformin enhances insulin binding to receptors on IM-9 cells and that this effect on insulin receptors may be related to alterations in metabolic functions that are reflected by a lowering of ATP levels.