Reactive centre loop mutants of α-1-antitrypsin reveal position-specific effects on intermediate formation along the polymerization pathway

The common severe Z mutation (E342K) of α₁-antitrypsin forms intracellular polymers that are associated with liver cirrhosis. The native fold of this protein is well-established and models have been proposed from crystallographic and biophysical data for the stable inter-molecular configuration that terminates the polymerization pathway. Despite these molecular ‘snapshots’, the details of the transition between monomer and polymer remain only partially understood. We surveyed the RCL (reactive centre loop) of α₁-antitrypsin to identify sites important for progression, through intermediate states, to polymer. Mutations at P₁₄P₁₂ and P₄, but not P₁₀P₈ or P₂P_1′, resulted in a decrease in detectable polymer in a cell model that recapitulates the intracellular polymerization of the Z variant, consistent with polymerization from a near-native conformation. We have developed a FRET (Förster resonance energy transfer)-based assay to monitor polymerization in small sample volumes. An in vitro assessment revealed the position-specific effects on the unimolecular and multimolecular phases of polymerization: the P₁₄P₁₂ region self-inserts early during activation, while the interaction between P₆P₄ and β-sheet A presents a kinetic barrier late in the polymerization pathway. Correspondingly, mutations at P₆P₄, but not P₁₄P₁₂, yield an increase in the overall apparent activation energy of association from ~360 to 550 kJ mol⁻¹.     

What happens to translocated game birds that 'disappear'?

The ultimate goal of most translocation efforts is to create a self-sustaining wild population of a species deliberately moved from one part of their range to another. As follow-up of a translocation attempt is often difficult, causes for failure are relatively unknown. Dispersal away from the release site is one potential source of failure because it decreases the likelihood of the released population establishing itself post-translocation. In this study, we used chukar Alectoris chukar as a surrogate for translocated game birds in order to conduct a large-scale experimental study. We observed that these desert-adapted birds demonstrate a strong fidelity for specific water sources. We also report the propensity for the translocated individuals to either disperse and return to their original water source site or remain at the release site. During two field seasons, we observed opposing behaviors such that the proportion of individuals returning to the capture site, versus those remaining at the release site, shifted between years. We analyzed this change between the years as well as within the years to assess the potential underlying causes such as translocated distance, differences in rainfall between seasons and water source type. We concluded that homing behavior was strong in this non-migratory bird species and that strength of this homing behavior varied, potentially due to conditions surrounding the limiting resource, water availability. The large-scale, original data presented here may help to explain why some releases result in a successfully established population while other releases result in widely dispersed individuals.     

Characterization of an Enantioselective Odorant Receptor in the Yellow Fever Mosquito Aedes aegypti

Enantiomers differ only in the left or right handedness (chirality) of their orientations and exhibit identical chemical and physical properties. In chemical communication systems, enantiomers can be differentially active at the physiological and behavioral levels. Only recently were enantioselective odorant receptors demonstrated in mammals while their existence in insects has remained hypothetical. Using the two-microelectrode voltage clamp of Xenopus oocytes, we show that the yellow fever mosquito, Aedes aegypti, odorant receptor 8 (AaOR8) acts as a chiral selective receptor for the (R)-(—)-enantiomer of 1-octen-3-ol, which in the presence of other kairomones is an attractant used by blood-sucking insects to locate their hosts. In addition to steric constraints, chain length and degree of unsaturation play important roles in this recognition process. This is the first characterization of an enantioselective odorant receptor in insects and the results demonstrate that an OR alone, without helper proteins, can account for chiral specificity exhibited by olfactory sensory neurons (OSNs).     

Combined effects of molt and chronic stress on heart rate, heart rate variability, and glucocorticoid physiology in European Starlings

Molt is an important life-history stage in avian species, but little is known about the effects of chronic stress during this period. Three weeks after the onset of molt, captive European starlings (Sturnus vulgaris) were exposed to 18 days of chronic stress, induced with four 30-minute randomized stressors presented daily. Birds showed no chronic-stress-induced changes in heart rate or heart rate variability when measured either during the middle of the day or at night. These data suggest that chronic stress did not alter the balance between sympathetic and parasympathetic nervous system regulation of cardiovascular function, which contrasts with data from an earlier study indicating that chronic stress profoundly alters cardiovascular function in non-molting starlings. Additionally, there was a significant increase in restraint-induced corticosterone secretion the first week of chronic stress that subsequently returned to pre-chronic-stress levels by the second week of exposure. The attenuated corticosterone response again contrasts with data from non-molting starlings that showed significant decreases in corticosterone responses. Consequently, the resistance to cardiovascular and corticosterone changes indicates that the physiological changes induced by chronic stress are greatly attenuated in molting birds. Overall, the data suggest that molt requires a degree of physiological stability that must be protected, so that if a bird is exposed to chronic stress during this life-history stage, molt takes priority.     

Calcium ions and osteoclastogenesis initiate the induction of bone formation by coral‐derived macroporous constructs

Coral-derived calcium carbonate/hydroxyapatite macroporous constructs of the genus Goniopora with limited hydrothermal conversion to hydroxyapatite (7% HA/CC) initiate the induction of bone formation. Which are the molecular signals that initiate pattern formation and the induction of bone formation? To evaluate the role of released calcium ions and osteoclastogenesis, 7% HA/CC was pre-loaded with either 500 μg of the calcium channel blocker, verapamil hydrochloride, or 240 μg of the osteoclast inhibitor, biphosphonate zoledronate, and implanted in the rectus abdominis muscle of six adult Chacma baboons Papio ursinus. Generated tissues on days 15, 60 and 90 were analysed by histomorphometry and qRT-PCR. On day 15, up-regulation of type IV collagen characterized all the implanted constructs correlating with vascular invasion. Zoledronate-treated specimens showed an important delay in tissue patterning and morphogenesis with limited bone formation. Osteoclastic inhibition yielded minimal, if any, bone formation by induction. 7% HA/CC pre-loaded with the Ca⁺⁺ channel blocker verapamil hydrochloride strongly inhibited the induction of bone formation. Down-regulation of bone morphogenetic protein-2 (BMP-2) together with up-regulation of Noggin genes correlated with limited bone formation in 7% HA/CC pre-loaded with either verapamil or zoledronate, indicating that the induction of bone formation by coral-derived macroporous constructs is via the BMPs pathway. The spontaneous induction of bone formation is initiated by a local peak of Ca⁺⁺ activating stem cell differentiation and the induction of bone formation.     

Olfactory Cues Are Subordinate to Visual Stimuli in a Neotropical Generalist Weevil

The tropical root weevil Diaprepes abbreviatus is a major pest of multiple crops in the Caribbean Islands and has become a serious constraint to citrus production in the United States. Recent work has identified host and conspecific volatiles that mediate host- and mate-finding by D. abbreviatus. The interaction of light, color, and odors has not been studied in this species. The responses of male and female D. abbreviatus to narrow bandwidths of visible light emitted by LEDs offered alone and in combination with olfactory stimuli were studied in a specially-designed multiple choice arena combined with a locomotion compensator. Weevils were more attracted to wavelengths close to green and yellow compared with blue or ultraviolet, but preferred red and darkness over green. Additionally, dim green light was preferred over brighter green. Adult weevils were also attracted to the odor of its citrus host + conspecifics. However, the attractiveness of citrus + conspecific odors disappeared in the presence of a green light. Photic stimulation induced males but not females to increase their speed. In the presence of light emitted by LEDs, turning speed decreased and path straightness increased, indicating that weevils tended to walk less tortuously. Diaprepes abbreviatus showed a hierarchy between chemo- and photo-taxis in the series of experiments presented herein, where the presence of the green light abolished upwind anemotaxis elicited by the pheromone + host plant odor. Insight into the strong responses to visual stimuli of chemically stimulated insects may be provided when the amount of information supplied by vision and olfaction is compared, as the information transmission capacity of compound eyes is estimated to be several orders of magnitude higher compared with the olfactory system. Subordination of olfactory responses by photic stimuli should be considered in the design of strategies aimed at management of such insects.