Influence of Cold on Radiostrontium Excretion

IN most work on the problems of releasing radiostrontium which has been incorporated into the vertebrate skeleton the effects of various parenterally administered chemical agents on the excretion rate of radiostrontium have been studied. Someriecrease in skeletal retention was obtained when inactive salts of strontium^1–4 or zirconium⁵ were given almost simultaneously with radiostrontium; so far all other efforts have been practically useless.     

The larval head of Nevrorthidae and the phylogeny of Neuroptera (Insecta)

External and internal head structures of larvae of Nevrorthidae were described in detail. The results were compared to conditions found in other representatives of Neuroptera and the other two neuropterid orders. The cladistic analysis supported the monophyly of Neuroptera, Neuroptera exclusive of Nevrorthidae, Hemerobiiformia, and Myrmeleontiformia. Neuroptera exclusive of Nevrorthidae are supported by the formation of an undivided postmentum and the presence of cryptonephric Malpighian tubules. The highly specialized articulation of the neck (Rollengelenk) and the absence of a salivary duct are autapomorphies of Nevrorthidae. Ithonidae and Polystoechotidae form a clade and are the sister group of the remaining Hemerobiiformia, which are characterized by the complete lack of a gula and a terminal filament of the antenna. Within this lineage, a clade comprising Mantispidae, Dilaridae, Berothidae, and Rhachiberothidae is well supported. Larvae of Myrmeleontiformia are characterized by a complex transformation of head structures, with a hypostomal bridge, a small triangular gula, largely reduced maxillary grooves, and anteriorly shifted posterior tentorial grooves. The slender finger‐like mid‐dorsal apodeme is another autapomorphy of the group. Psychopsidae are placed as the sister group of the remaining Myrmeleontiformia, which are characterized by a conspicuous, protruding ocular region (often less distinct or even absent in Nemopteridae). Ascalaphidae are the sister group of Myrmeleontidae. Larvae of both families share the fusion of the tibia and tarsus in the hind leg. The larval characters analysed were not sufficient for full resolution of the myrmeleontiform and hemerobiiform lineages. The position of several families such as Osmylidae, Sisyridae, and Coniopterygidae remains uncertain. The results are in agreement with an aquatic ancestor of Neuroptera and secondarily acquired terrestrial habits within the lineage (Neuroptera exclusive of Nevrorthidae), and another invasion of the aquatic environment by Sisyridae. © 2010 The Linnean Society of London, Zoological Journal of the Linnean Society, 2010, 158 , 533–562.     

Monocyte lipid rafts contain proteins implicated in vesicular trafficking and phagosome formation

Lipid rafts are membrane microdomains of unique lipid composition that segregate proteins with poorly understood consequences for membrane organization. Identification of raft associated proteins could therefore provide novel insight into raft-dependent functions. Monocytes process antigens for presentation to T cells by ingesting pathogens into calcium-dependent plasma membrane invaginations called "phagosomes" which develop by sequential fusion with the endoplasmic reticulum, early and late endosomes. We investigated the protein composition of Triton X-100 insoluble low density membranes of the monocyte cell-line THP-1 by matrix-assisted laser desorption/ionization-time of flight and tandem mass spectrometry. The ganglioside GM1 colocalized on the plasma membrane with the raft markers flotillin 1 and 2, which were enriched in low buoyant density fractions containing 52 identifiable proteins, 28 of which have not been reported in rafts, and nine of which are associated with the endoplasmic reticulum (ER). Remarkably, 27 of the 52 proteins are components of phagosomes, including the ER protein calnexin which we demonstrate is phosphorylated on serine 562, a switch controlling calcium homeostasis. The presence of the early and late endosome trafficking proteins Rab-1, and Rab-7 together with the late endosome protein LIMPII, indicate lipid rafts are present throughout endosome maturation. Identification of vacuolar ATP synthase, and synaptosomal-associated protein-23, proteins implicated in membrane fusion, together with the cytoskeletal proteins actin, alpha-actinin, and vimentin, and Rac 1, 2, and 3, regulators of cytoskeletal assembly, indicate monocyte lipid rafts contain the machinery to direct vesicular fusion and actin based vesicular migration throughout phagosome development.     

Winter herbivory by voles during a population peak: the relative importance of local factors and landscape pattern

1. We studied the relative role of local habitat variables and landscape pattern on vole–plant interactions in a system with grey-sided voles ( Clethrionomys rufocanus (Sund.)) and their favourite winter food plant, bilberry ( Vaccinium myrtillus L.). The study was conducted during a vole peak year (1992–93) in a tundra area in northern Norway.
2. Using Mantel statistics we were able to separate the direct effects of the spatial patterning of habitats and the indirect effects due to spatial aggregations of similar habitats.
3. Results indicate that knowledge about the explicit spatial patterning of patches does not improve our understanding of the system. Instead, two local factors, vegetation height and bilberry biomass, explained more than 50% of the variation in cutting intensity in winter (defined as the proportion of above-ground shoots cut). Increasing vegetation height increased, and increasing bilberry biomass decreased, the cutting intensity.
4. The conclusion that grey-sided voles are able to distribute themselves relative to habitat quality was also partially supported by our estimated over-winter persistence by voles in the various habitats. Vole persistence was uncorrelated with vegetation height, the important predictor of autumn vole density, but tended to correlate with the deviation from the relation between vegetation height and autumn vole density. This conforms to the expectations from the theory of ideal-free habitat distribution.
5. The cue for vole habitat choice, i.e. vegetation height, indicates that either predation or freezing risk is important for voles when selecting over-wintering habitat.     

High misses after odd flashes in oxygen evolution in thoroughly dark-adapted thylakoids from pea and Chenopodium album

In Photosystem II (PS II), water is oxidized to molecular oxygen and plastoquinone is reduced to plastoquinol. The oxidation of water requires the accumulation of four oxidizing equivalents, through the so-called S-states of the oxygen evolving complex; the production of plastoquinol requires the accumulation of two reducing equivalents on a bound plastoquinone, Q_B. It has been generally believed that during the flash-induced transition of each of the S-states (S_n S_n+1, where n=0, 1, 2 and 3), a certain small but equal fraction of the PS II reaction centers are unable to function and, thus, miss being turned over. We used thoroughly dark-adapted thylakoids from peas (Pisum sativum) and Chenopodium album (susceptible and resistant to atrazine) starting with 100% of the oxygen evolving complex in the S₁ state. Thylakoids were illuminated with saturating flashes, providing a double hit parameter of about 0.07. Our experimental data on flashnumber dependent oscillations in the amount of oxygen per flash fit very well with a binary pattern of misses: 0, 0.2, 0, 0.4 during S₀ S₁, S₁ S₂, S₂ S₃ and S₃ S₀ transitions. Addition of 2 mM ferricyanide appears to shift this pattern by one flash. These results are consistent with the bicycle model recently proposed by V. P. Shinkarev and C. A. Wraight (Oxygen evolution in photosynthesis: From unicycle to bicycle, 1993, Proc Natl Acad Sci USA 90: 1834–1838), where misses are due to the presence of P⁺ or Q_A ^- among the various equilibrium states of PS II centers.Abbreviations miss parameter - double hit parameter - PS II Photosystem II - Q_A primary one-electron acceptor of PS II, a plastoquinone molecule - Q_B secondary plastoquinone two-electron acceptor of PS II - S-states (S_n, where n=0, 1, 2, 3 or 4) redox states of the oxygen evolving complex     

Insect crossbridges, relaxed by spin-labeled nucleotide, show well-ordered 90 degrees state by X-ray diffraction and electron microscopy, but spectra of electron paramagnetic resonance probes report disorder.

The structure of glycerinated Lethocerus insect flight muscle fibers, relaxed by spin-labeled ATP and vanadate (Vi), was examined using X-ray diffraction, electron microscopy and electron paramagnetic resonance (e.p.r.) spectra. We obtained excellent relaxation of MgATP quality as determined by mechanical criteria, using vanadate trapping of 2' spin-labeled 3' deoxyATP at 3 degree C. In rigor fibers, when the diphosphate analog is bound in the absence of Vi, the probes on myosin heads are well-ordered, in agreement with electron microscopic and X-ray patterns showing that myosin heads are ordered when attached strongly to actin. In relaxed muscle, however, e.p.r. spectra report orientational disorder of bound (Vi-trapped) spin-labeled nucleotide, while electron microscopic and X-ray patterns both show well-ordered bridges at a uniform 90 degrees angle to the filament axis. The spin-labeled nucleotide orientation is highly disordered, but not completely isotropic; the slight anisotropy observed in probe spectra is consistent with a shift of approximately 10% of probes from angles close to 0 degrees to angles close to 90 degrees. Measurements of probe mobility suggest that the interaction between probe and protein remains as tight in relaxed fibers as in rigor, and thus that the disorder in relaxed fibers arises from disorders of (or within) the protein and not from disorder of the probe relative to the protein. Fixation of the relaxed fibers with glutaraldehyde did not alter any aspect of the spectrum of the Vi-trapped analog, including the slight order observed, showing that the extensive inter- and intra-molecular cross-linking of the first step of sample preparation for electron microscopy had not altered relaxed crossbridge orientations. Two models that may reconcile the apparently disparate results obtained on relaxed fibers are presented: (1) a rigid myosin head could possess considerable disorder in the regular array about the thick filament; or (2) the nucleotide site could be on a disordered, probably distal, domain of myosin, while a more proximal region is well ordered on the thick filament backbone. Our findings suggest that when e.p.r. probes signal disorder of a local site or domain, this is complementary, not contradictory, to signals of general order. The e.p.r. spectra show that a portion of the myosin molecule can be disordered at the same time as the X-ray diffraction and electron microscopy show the bulk of myosin head mass to be uniformly oriented and regularly arrayed.     

The Kinesin-1 Tail Conformationally Restricts the Nucleotide Pocket

We have used electron paramagnetic resonance and fluorescence spectroscopy to study the interaction between the kinesin-1 head and its regulatory tail domain. The interaction between the tails and the enzymatically active heads has been shown to inhibit intrinsic and microtubule-stimulated ADP release. Here, we demonstrate that the probe mobility of two different spin-labeled nucleotide analogs in the kinesin-1 nucleotide pocket is restricted upon binding of the tail domain to kinesin-1 heads. This conformational restriction is distinct from the microtubule-induced changes in the nucleotide pocket. Unlike myosin V, this tail-induced restriction occurs independent of nucleotide state. We find that the head-tail interaction that causes the restriction only weakly stabilizes Mg²⁺ in the nucleotide pocket. The conformational restriction also occurs when a tail construct containing a K922A point mutation is used. This mutation eliminates the tail's ability to inhibit ADP release, indicating that the tail does not inhibit nucleotide ejection from the pocket by simple steric hindrance. Together, our data suggest that the observed head-tail interaction serves as a scaffold to position K922 to exert its inhibitory effect, possibly by interacting with the nucleotide α/β-phosphates in a manner analogous to the arginine finger regulators of some G proteins.     

The Histone H4 Tail Regulates the Conformation of the ATP-Binding Pocket in the SNF2h Chromatin Remodeling Enzyme

The chromatin remodeling complex ACF helps establish the appropriate nucleosome spacing for generating repressed chromatin states. ACF activity is stimulated by two defining features of the nucleosomal substrate: a basic patch on the histone H4 N-terminal tail and the specific length of flanking DNA. However, the mechanisms by which these two substrate cues function in the ACF remodeling reaction is not well understood. Using electron paramagnetic resonance spectroscopy with spin-labeled ATP analogs to probe the structure of the ATP active site under physiological solution conditions, we identify a closed state of the ATP-binding pocket that correlates with ATPase activity. We find that the H4 tail promotes pocket closure. We further show that ATPase stimulation by the H4 tail does not require a specific structure connecting the H4 tail and the globular domain. In the case of many DNA helicases, closure of the ATP-binding pocket is regulated by specific DNA substrates. Pocket closure by the H4 tail may analogously provide a mechanism to directly couple substrate recognition to activity. Surprisingly, the flanking DNA, which also stimulates ATP hydrolysis, does not promote pocket closure, suggesting that the H4 tail and flanking DNA may be recognized in different reaction steps.     

High-resolution near-field optical imaging of single nuclear pore complexes under physiological conditions

Scanning near-field optical microscopy (SNOM) circumvents the diffraction limit of conventional light microscopy and is able to achieve optical resolutions substantially below 100 nm. However, in the field of cell biology SNOM has been rarely applied, probably because previous techniques for sample-distance control are less sensitive in liquid than in air. Recently we developed a distance control based on a tuning fork in tapping mode, which is also well-suited for imaging in solution. Here we show that this approach can be used to visualize single membrane protein complexes kept in physiological media throughout. Nuclear envelopes were isolated from Xenopus laevis oocytes at conditions shown recently to conserve the transport functions of the nuclear pore complex (NPC). Isolated nuclear envelopes were fluorescently labeled by antibodies against specific proteins of the NPC (NUP153 and p62) and imaged at a resolution of approximately 60 nm. The lateral distribution of epitopes within the supramolecular NPC could be inferred from an analysis of the intensity distribution of the fluorescence spots. The different number densities of p62- and NUP153-labeled NPCs are determined and discussed. Thus we show that SNOM opens up new possibilities for directly visualizing the transport of single particles through single NPCs and other transporters.