The effect of branch lengths on phylogeny: an empirical study using highly conserved orthologs from mammalian genomes

Phylogenetic analyses were applied to 269 families of putative orthologs represented by a single member in the genomes of human, mouse, dog, and chicken. Five methods were used: maximum parsimony (NP), neighbor-joining (NJ) with Poisson and Gamma distances; and maximum likelihood (ML) with JTT and JTT+gamma models. When applied to the concatenated sequence of all families, all methods strongly supported a tree in which mouse branched before human and dog. In analyses of individual families, the same topology was supported more than any other. Although there was evidence of an increased rate of amino acid replacement in the mouse lineage in comparison to the other two mammals, there was no evidence that support for the mouse's basal position was due to long-branch attraction; rather, this topology was seen in the families with the lowest rate variation among the three mammalian branches. In families with highly divergent mouse sequences, ML with both JTT and JTT+gamma and NJ with the gamma distance tended to support a topology in which the dog, rather than the mouse, branched first. Thus, in these data, a tendency of long and short branches to cluster together ("opposite-branch attraction") seemed to be more of a problem than long-branch attraction.     

Likelihood-ratio tests for positive selection of human and mouse duplicate genes reveal nonconservative and anomalous properties of widely used methods

Two commonly used methods based on likelihood-ratio tests (LRTs) for detecting positive Darwinian selection at the molecular level were applied to a data set of 604 gene families containing two members in the human genome and two members in the mouse genome. These methods detected positive selection in a very high proportion of families; in over 50% of families, there was significant evidence of positive selection by one or both methods. However, less than a third of families showing evidence for positive selection by at least one of the methods showed evidence of positive selection by both methods. The outcome of these tests was predicted better by sequence length, G+C content at third-codon positions, and the level of synonymous substitution than by the level of nonsynonymous substitution or the ratio of nonsynonymous to synonymous substitution. These results suggested that LRT-based tests for positive selection may be sensitive to certain factors that make it difficult to reconstruct the true pattern of nucleotide substitution.     

Flight responses of eastern gray kangaroos to benign or harmful human behavior

Globally, wilderness is being converted for rural and agricultural land use. In countryside landscapes, many habitat structures remain intact, providing suitable habitat for wildlife species that can accurately assess novel risks and develop tolerance to benign disturbances. Associative learning that promotes avoidance and also facilitates desensitization to benign disturbance is key to persisting in these landscapes. Conversely, learning to distinguish and avoid negative interactions with humans, like hunting, is vital. To determine if eastern gray kangaroos are capable of learning from previous interactions with humans, we tested the flight responses of wild kangaroos which have previously experienced either low or high frequencies of harmful and benign encounters with humans. We found that eastern gray kangaroos rapidly habituated to benign disturbance as there was no significant difference in assessment distance between groups that previously experienced low or high frequencies of disturbance. The threat of harmful disturbances was not as quickly learnt, as groups that experienced low frequencies of harmful disturbance delayed flight longer than those experiencing frequent harm. We found that the influence of environmental and group parameters on a kangaroo's decision to flee depended on the intent and frequency of previous interactions with humans. Our study indicates that kangaroos are learning from previous encounters with humans, correctly assessing novel risks which may be contributing to their persistence in countryside landscapes.     

Control of Insulin Secretion by Cytochrome c and Calcium Signaling in Islets with Impaired Metabolism

The aim of the study was to assess the relative control of insulin secretion rate (ISR) by calcium influx and signaling from cytochrome c in islets where, as in diabetes, the metabolic pathways are impaired. This was achieved either by culturing isolated islets at low (3 mm) glucose or by fasting rats prior to the isolation of the islets. Culture in low glucose greatly reduced the glucose response of cytochrome c reduction and translocation and ISR, but did not affect the response to the mitochondrial fuel α-ketoisocaproate. Unexpectedly, glucose-stimulated calcium influx was only slightly reduced in low glucose-cultured islets and was not responsible for the impairment in glucose-stimulated ISR. A glucokinase activator acutely restored cytochrome c reduction and translocation and ISR, independent of effects on calcium influx. Islets from fasted rats had reduced ISR and cytochrome c reduction in response to both glucose and α-ketoisocaproate despite normal responses of calcium. Our data are consistent with the scenario where cytochrome c reduction and translocation are essential signals in the stimulation of ISR, the loss of which can result in impaired ISR even when calcium response is normal.     

Small Substrate Transport and Mechanism of a Molybdate ATP Binding Cassette Transporter in a Lipid Environment

Embedded in the plasma membrane of all bacteria, ATP binding cassette (ABC) importers facilitate the uptake of several vital nutrients and cofactors. The ABC transporter, MolBC-A, imports molybdate by passing substrate from the binding protein MolA to a membrane-spanning translocation pathway of MolB. To understand the mechanism of transport in the biological membrane as a whole, the effects of the lipid bilayer on transport needed to be addressed. Continuous wave-electron paramagnetic resonance and in vivo molybdate uptake studies were used to test the impact of the lipid environment on the mechanism and function of MolBC-A. Working with the bacterium Haemophilus influenzae, we found that MolBC-A functions as a low affinity molybdate transporter in its native environment. In periods of high extracellular molybdate concentration, H. influenzae makes use of parallel molybdate transport systems (MolBC-A and ModBC-A) to take up a greater amount of molybdate than a strain with ModBC-A alone. In addition, the movement of the translocation pathway in response to nucleotide binding and hydrolysis in a lipid environment is conserved when compared with in-detergent analysis. However, electron paramagnetic resonance spectroscopy indicates that a lipid environment restricts the flexibility of the MolBC translocation pathway. By combining continuous wave-electron paramagnetic resonance spectroscopy and substrate uptake studies, we reveal details of molybdate transport and the logistics of uptake systems that employ multiple transporters for the same substrate, offering insight into the mechanisms of nutrient uptake in bacteria.