Impaired pulmonary host defense in mice lacking expression of the CXC chemokine lungkine

Lungkine (CXCL15) is a novel CXC chemokine that is highly expressed in the adult mouse lung. To determine the biologic function of Lungkine, we generated Lungkine null mice by targeted gene disruption. These mice did not differ from wild-type mice in their hematocrits or in the relative number of cells in leukocyte populations of peripheral blood or other tissues, including lung and bone marrow. However, Lungkine null mice were more susceptible to Klebsiella pneumonia infection, with a decreased survival and increased lung bacterial burden compared with infected wild-type mice. Histologic analysis of the lung and assessment of leukocytes in the bronchioalveolar lavage revealed that neutrophil numbers were normal in the lung parenchyma, but reduced in the airspace. The production of other neutrophil chemoattractants in the Lungkine null mice did not differ from that in wild-type mice, and neutrophil migration into other tissues was normal. Taken together, these findings demonstrate that Lungkine is an important mediator of neutrophil migration from the lung parenchyma into the airspace.     

Dynamic regimes and correlated structural dynamics in native and denatured alpha-lactalbumin

Understanding the mechanisms of protein folding requires knowledge of both the energy landscape and the structural dynamics of a protein. We report a neutron-scattering study of the nanosecond and picosecond dynamics of native and the denatured alpha-lactalbumin. The quasielastic scattering intensity shows that there are alpha-helical structure and tertiary-like side-chain interactions fluctuating on sub-nanosecond time-scales under extremely denaturing conditions and even in the absence of disulfide bonds. Based on the length-scale dependence of the decay rate of the measured correlation functions, the nanosecond dynamics of the native and the variously denatured proteins have three dynamic regimes. When 0.051.0 A(-1) is a regime that displays the local dynamic behavior of individual residues, Gamma proportional to Q(1.8+/-0.3). The picosecond time-scale dynamics shows that the potential barrier to side-chain proton jump motion is reduced in the molten globule and in the denatured proteins when compared to that of the native protein. Our results provide a dynamic view of the native-like topology established in the early stages of protein folding.     

Spatial variability in oviposition damage by periodical cicadas in a fragmented landscape

Effects of the periodical cicada (Magicicada spp.) on forest dynamics are poorly documented. A 1998 emergence of M. cassini in eastern Kansas led to colonization of a fragmented experimental landscape undergoing secondary succession. We hypothesized that per-tree rates of oviposition damage by cicadas would reflect: (1) distance from the source of the emergence, (2) patch size, and (3) local tree density. Ovipositing females displayed clear preferences for host species and damage incidence showed predictable spatial patterns. Two species (smooth sumac, Rhus glabra, and eastern red cedar, Juniperus virginiana) were rarely attacked, whereas others (rough-leaved dogwood, Cornus drummondii; slippery elm, Ulmus rubra; box elder, Acer negundo, and honey locust, Gleditsia triacanthos) were strongly attacked. The dominant early successional tree, dogwood, received on average the most attacks. As predicted, attacks per stem declined strongly with distance from the emergence source, and with local stem density (a "dilution" effect). Contrary to expectations, there were more attacks per stem on larger patches. Because ovipositing cicadas cut damaging slits in host tree branches, potentially affecting tree growth rate, competitive ability, and capacity to reproduce, cicada damage could potentially influence spatial variation in secondary succession.     

A mitochondrial control region and cytochrome b phylogeny of sika deer (Cervus nippon) and report of tandem repeats in the control region

Sika deer (Cervus nippon Temminck) are endemic to mainland and insular Asia. Numerous subspecies have been named, but they are not quantitatively well defined. Portions of the mitochondrial cytochrome b gene (450 bp) and control region (512 bp) were sequenced from 28 individuals belonging to five sika subspecies and two Cervus elaphus subspecies. Phylogenetic trees constructed using these sequences clearly demonstrated that sika are monophyletic with respect to C. elaphus. A survey of variation in the control region showed that approximately half the variation occurred in a 100-base segment between positions 150 and 250 in the left domain of the control region. Within this region there were three tandemly repeated copies of a 39-base motif. In addition, two of the samples (C. n. aplodontus and C. n. hortulorum) contained, respectively, two and four additional copies of the repeated motif.     

Microtubules and signal transduction

Although molecular components of signal transduction pathways are rapidly being identified, how elements of these pathways are positioned spatially and how signals traverse the intracellular environment from the cell surface to the nucleus or to other cytoplasmic targets are not well understood. The discovery of signaling molecules that interact with microtubules (MTs), as well as the multiple effects on signaling pathways of drugs that destabilize or hyperstabilize MTs, indicate that MTs are likely to be critical to the spatial organization of signal transduction. MTs themselves are also affected by signaling pathways and this may contribute to the transmission of signals to downstream targets.     

The transmission and effects of Wolbachia bacteria in parasitoids

Wolbachia bacteria are obligatory intracellular parasites of arthropods and have been detected in about 70 species of parasitic wasps and three parasitoid flies. Wolbachia are transmitted cytoplasmically (maternally) and modify host reproduction in different ways to enhance their own transmission: parthenogenesis induction (PI), cytoplasmic incompatibility (CI), or feminization (F) of genetic males. Only PI and CI are known in parasitoids. PI-Wolbachia cause thelytoky in otherwise arrhenotokous parasitoids by generating diploid (rather than haploid) unfertilized wasp eggs. CI-Wolbachia cause incompatibility of crosses between infected males and uninfected females because the paternally derived chromosomes fail to decondense and are destroyed after syngamy. More complex situations arise when hosts harbor multiple infections, which can lead to bidirectional incompatibility and may be involved in parasitoid speciation. The relative fitness of infected and uninfected hosts is important to the population dynamics of Wolbachia, and more data are needed. Evolutionary conflict should be common between host genes, Wolbachia genes, and other "selfish" genetic elements. Wolbachia-specific PCR primers are now available for several genes with different rates of evolution. These primers will permit rapid screening in future studies of spatial and temporal patterns of single and multiple infection. Molecular phylogenies show that CI- and PI-Wolbachia do not form discrete clades. In combination with experimental transfection data, this result suggests that host reproductive alterations depend on the interaction between attributes of both Wolbachia and host. Moreover, Wolbachia isolates from closely related hosts do not usually cluster together, and phylogenies suggest that Wolbachia may have radiated after their arthropod hosts. Both results support considerable horizontal transmission of Wolbachia between host species over evolutionary time. Natural horizontal transmisson between parasitoids and their hosts, or with entomoparasitic nematodes or ectoparasitic mites, remains a tantalizing but equivocal possibility. Received: November 27, 1998 / Accepted: January 15, 1999     

Direct imaging of DNA in living cells reveals the dynamics of chromosome formation

Individual chromosomes are not directly visible within the interphase nuclei of most somatic cells; they can only be seen during mitosis. We have developed a method that allows DNA strands to be observed directly in living cells, and we use it to analyze how mitotic chromosomes form. A fluorescent analogue (e.g., Cy5-dUTP) of the natural precursor, thymidine triphosphate, is introduced into cells, which are then grown on the heated stage of a confocal microscope. The analogue is incorporated by the endogenous enzymes into DNA. As the mechanisms for recognizing and removing the unusual residues do not prevent subsequent progress around the cell cycle, the now fluorescent DNA strands can be followed as they assemble into chromosomes, and segregate to daughters and granddaughters. Movies of such strands in living cells suggest that chromosome axes follow simple recognizable paths through their territories during G2 phase, and that late replicating regions maintain their relative positions as prophase chromosomes form. Quantitative analysis confirms that individual regions move little during this stage of chromosome condensation. As a result, the gross structure of an interphase chromosome territory is directly related to that of the prophase chromosome.     

The L1 major capsid protein of human papillomavirus type 11 recombinant virus-like particles interacts with heparin and cell-surface glycosaminoglycans on human keratinocytes

The L1 major capsid protein of human papillomavirus (HPV) type 11, a 55-kDa polypeptide, forms particulate structures resembling native virus with an average particle diameter of 50-60 nm when expressed in the yeast Saccharomyces cerevisiae. We show in this report that these virus-like particles (VLPs) interact with heparin and with cell-surface glycosaminoglycans (GAGs) resembling heparin on keratinocytes and Chinese hamster ovary cells. The binding of VLPs to heparin is shown to exhibit an affinity comparable to that of other identified heparin-binding proteins. Immobilized heparin chromatography and surface plasmon resonance were used to show that this interaction can be specifically inhibited by free heparin and dextran sulfate and that the effectiveness of the inhibitor is related to its molecular weight and charge density. Sequence comparison of nine human L1 types revealed a conserved region of the carboxyl terminus containing clustered basic amino acids that bear resemblance to proposed heparin-binding motifs in unrelated proteins. Specific enzymatic cleavage of this region eliminated binding to both immobilized heparin and human keratinocyte (HaCaT) cells. Removal of heparan sulfate GAGs on keratinocytes by treatment with heparinase or heparitinase resulted in an 80-90% reduction of VLP binding, whereas treatment of cells with laminin, a substrate for alpha6 integrin receptors, provided minimal inhibition. Cells treated with chlorate or substituted beta-D-xylosides, resulting in undersulfation or secretion of GAG chains, also showed a reduced affinity for VLPs. Similarly, binding of VLPs to a Chinese hamster ovary cell mutant deficient in GAG synthesis was shown to be only 10% that observed for wild type cells. This report establishes for the first time that the carboxyl-terminal portion of HPV L1 interacts with heparin, and that this region appears to be crucial for interaction with the cell surface.