Activity-dependent α-Cleavage of Nectin-1 Is Mediated by A Disintegrin and Metalloprotease 10 (ADAM10)

Nectin-1 is known to undergo ectodomain shedding by α-secretase and subsequent proteolytic processing by γ-secretase. How secretase-mediated cleavage of nectin-1 is regulated in neuronal cells and how nectin-1 cleavage affects synaptic adhesion is poorly understood. We have investigated α-and γ-secretase-mediated processing of nectin-1 in primary cortical neurons and identified which protease acts as a α-secretase. We report here that NMDA receptor activation, but not stimulation of AMPA or metabotropic glutamate receptors, resulted in robust α- and γ-secretase cleavage of nectin-1 in mature cortical neurons. Cleavage of nectin-1 required influx of Ca²⁺ through the NMDA receptor, and activation of calmodulin, but was not dependent on calcium/calmodulin-dependent protein kinase II (CaMKII) activation. We found that ADAM10 is the major secretase responsible for nectin-1 ectodomain cleavage in neurons and the brain. These observations suggest that α- and γ-secretase processing of nectin-1 is a Ca²⁺/calmodulin-regulated event that occurs under conditions of activity-dependent synaptic plasticity and ADAM10 and γ-secretase are responsible for these cleavage events.     

Mating Behavior of Cephalonomia tarsalis (Ashmead) (Hymenoptera: Bethylidae) and the Effect of Female Mating Frequency on Offspring Production

The courtship behavior of Cephalonomia tarsalis, a solitary semiectoparasitoid of Oryzaephilus surinamensis, was investigated in the laboratory. Courtship behavior includes a series of stereotypic movements. Males play the most active role, executing the majority of courtship action, and females respond with relatively limited observable behaviors. Males typically keep antennae still during encounters with females prior to mounting, which may be correlated with recognition of the female's sexual status. After mounting, males display a series of movements on females, such as antennae touching female's antennae, antennae or mouth touching female's head or thorax, and walking around on female, which may serve to stimulate females towards increased receptivity. Females signal receptivity by assuming a stereotypical posture of remaining stationary, with head down, and antennae still in front of the body. The male then inserts his aedeagus and the pair copulates. After an average of 40.4 s of copulation, females signal the end of copulation by waving the antennae and moving away from the copulation site. Males continue copulating for a short time after females start moving but dismount soon thereafter. After dismounting, the two wasps move away from each other immediately, and they typically begin grooming. Neither males nor females exhibit mating preference based on mate's mating status in both choice and no-choice tests. The male is polygynous and the mated female can mate multiple times within the first 3 days after starting oviposition. However, female mating frequency does not affect the production of female progeny.     

No neural evidence for dynamic auditory tuning of the A1 receptor in the ear of the noctuid moth, <Emphasis Type="Italic">Noctua pronuba</Emphasis>

By examining the mechanical properties of the tympanum of the noctuid moth, Noctua pronuba, Windmill et al. (2006) suggested that this insect increases (up-tunes) the frequencies of its best hearing when exposed to high intensity sounds (HIS) resembling the echolocation calls of attacking bats. We tested whether this biophysical phenomenon was encoded in the neural responses of this moth’s most sensitive auditory receptor (A1 cell) before and after exposure to HIS. We measured: (1) the number of A1 action potentials (spikes) per stimulus pulse; (2) the proportion of A1 spike periods below that determined to elicit evasive flight maneuvers and, (3) the change in A1 cell firing (spike number, interspike interval, stimulus/spike latency) over a duration of time similar to that in which up-tuning lasts. We observed no significant spiking response changes in the predicted direction to any of the frequencies tested following exposure to HIS and we observed only two of the 24 predicted time-dependent changes to A1 firing. These results indicate that tympanal up-tuning does not result in a change to this moth’s auditory frequency sensitivity and we suggest either sensillar resonances or increases in thoracic muscle tension following exposure to HIS as alternative explanations.     

Phospholamban C-terminal Residues Are Critical Determinants of the Structure and Function of the Calcium ATPase Regulatory Complex

To determine the structural and regulatory role of the C-terminal residues of phospholamban (PLB) in the membranes of living cells, we fused fluorescent protein tags to PLB and sarco/endoplasmic reticulum calcium ATPase (SERCA). Alanine substitution of PLB C-terminal residues significantly altered fluorescence resonance energy transfer (FRET) from PLB to PLB and SERCA to PLB, suggesting a change in quaternary conformation of PLB pentamer and SERCA-PLB regulatory complex. Val to Ala substitution at position 49 (V49A) had particularly large effects on PLB pentamer structure and PLB-SERCA regulatory complex conformation, increasing and decreasing probe separation distance, respectively. We also quantified a decrease in oligomerization affinity, an increase in binding affinity of V49A-PLB for SERCA, and a gain of inhibitory function as quantified by calcium-dependent ATPase activity. Notably, deletion of only a few C-terminal residues resulted in significant loss of PLB membrane anchoring and mislocalization to the cytoplasm and nucleus. C-terminal truncations also resulted in progressive loss of PLB-PLB FRET due to a decrease in the apparent affinity of PLB oligomerization. We quantified a similar decrease in the binding affinity of truncated PLB for SERCA and loss of inhibitory potency. However, despite decreased SERCA-PLB binding, intermolecular FRET for Val⁴⁹-stop (V49X) truncation mutant was paradoxically increased as a result of an 11.3-Å decrease in the distance between donor and acceptor fluorophores. We conclude that PLB C-terminal residues are critical for localization, oligomerization, and regulatory function. In particular, the PLB C terminus is an important determinant of the quaternary structure of the SERCA regulatory complex.     

Three-Dimensional Organization of Troponin on Cardiac Muscle Thin Filaments in the Relaxed State

Muscle contraction is regulated by troponin-tropomyosin, which blocks and unblocks myosin binding sites on actin. To elucidate this regulatory mechanism, the three-dimensional organization of troponin and tropomyosin on the thin filament must be determined. Although tropomyosin is well defined in electron microscopy helical reconstructions of thin filaments, troponin density is mostly lost. Here, we determined troponin organization on native relaxed cardiac muscle thin filaments by applying single particle reconstruction procedures to negatively stained specimens. Multiple reference models led to the same final structure, indicating absence of model bias in the procedure. The new reconstructions clearly showed F-actin, tropomyosin, and troponin densities. At the 25 Å resolution achieved, troponin was considerably better defined than in previous reconstructions. The troponin density closely resembled the shape of troponin crystallographic structures, facilitating detailed interpretation of the electron microscopy density map. The orientation of troponin-T and the troponin core domain established troponin polarity. Density attributable to the troponin-I mobile regulatory domain was positioned where it could hold tropomyosin in its blocking position on actin, thus suggesting the underlying structural basis of thin filament regulation. Our previous understanding of thin filament regulation had been limited to known movements of tropomyosin that sterically block and unblock myosin binding sites on actin. We now show how troponin, the Ca²⁺ sensor, may control these movements, ultimately determining whether muscle contracts or relaxes.     

Sex-biased natal dispersal and inbreeding avoidance in American black bears as revealed by spatial genetic analyses

We tested the hypothesis that sex-biased natal dispersal reduces close inbreeding in American black bears, a solitary species that exhibits nearly complete male dispersal and female philopatry. Using microsatellite DNA and spatial data from reproductively mature bears (>or= 4 years old), we examined the spatial genetic structure of two distinct populations in New Mexico from 1993 to 2000. As predicted, relatedness (r) and the frequency of close relationships (parent-offspring or full siblings) decreased with distance among female dyads, but little change was observed among male or opposite-sex dyads. Neighbouring females were more closely related than neighbouring males. The potential for inbreeding was low. Most opposite-sex pairs that lived sufficiently close to facilitate mating were unrelated, and few were close relatives. We found no evidence that bears actively avoided inbreeding in their selection of mates from this nearby pool, as mean r and relationship frequencies did not differ between potential and actual mating pairs (determined by parentage analysis). These basic patterns were apparent in both study areas despite a nearly two-fold difference in density. However, the sex bias in dispersal was less pronounced in the lower-density area, based on proportions of bears with male and female relatives residing nearby. This result suggests that male bears may respond to reduced competition by decreasing their rate or distance of dispersal. Evidence supports the hypothesis that inbreeding avoidance is achieved by means of male-biased dispersal but also indicates that competition (for mates or resources) modifies dispersal patterns.     

The structure of (μ3-sulfido)(μ3-p-tolyphosphido)tris(tricarbonyliron)(2FeFe)

Compounds of the type (μ₃-S)(μ₃-RP)Fe₃(CO)₉ have been prepared by the reaction of Fe₃(CO)₁₂ with [RP(S)S]₂ or RP(S)Cl₃. In this paper we report the synthesis of (μ₃-S)(μ₃-CH₃OC₆H₄P)Fe₃(CO)₉ using Lawesson's reagents, and the three dimensional structure of (μ₃-S)(μ₃-p-CH₃C₆H₄P)Fe₃(CO)₉. This material crystallizes in the space group P2₁/n with a = 8.558(2) Å, b = 9.022(2) Å, c = 27.506(6) Å, β = 95.40(2)°, Z = 4. The cluster is a nido structure as found commonly for (μ₃-X)(μ₃-Y)Fe₃(CO)₉ complexes.     

Mechanisms of acquired immunity in leishmaniasis

Self-curing cutaneous leishmaniasis depends on T cell-mediated immune activation of infected macrophages. Failure of immune control in inbred mouse models of metastasizing mucocutaneous and visceralizing forms of the disease involves, respectively, insusceptibility of the parasite and the generation of T cells that suppress a potentially curative response. Prophylactic immunization in man has so far been restricted to cutaneous leishmaniasis and based on inducing infection under controlled conditions with virulent Leishmania tropica major promastigotes. The feasibility of immunization against visceral leishmaniasis merits reconsideration. BALB/c mice are genetically vulnerable to L. tropica major, which produces a fatal visceralizing type of disease involving specific suppression of cell-mediated immunity. Potent and lasting protection can be induced by repeated intravenous immunization with irradiated promastigotes. The efficacy of this 'vaccine' is relatively heat-stable (1 h at 56 degrees C). Immunity is not attributable to antibody but to the generation of Lyt-1+2- T cells which, although possessing helper and macrophage-activating functions, do not express classical delayed-type hypersensitivity. The immunological features of this system and its relevance to the possibility of protection against human Leishmania donovani infection are considered.