Single anion channels reconstituted from cardiac mitoplasts

Ion channels from sheep cardiac mitoplast (inverted inner mitochondrial membrane vesicle) preparations were incorporated into voltage-clamped planar lipid bilayers. The appearance of anion rather than cation channels could be promoted by exposing the bilayers to osmotic gradients formed by Cl⁻ salts of large, relatively imperment, cations at a pH of 8.8. Two distinct activities were identified. These comprised a multisubstate anion channel of intermediate conductance (∼60 pS in 300vs. 50mm choline Cl, ∼100 pS in symmetric 150mm KCl), and a lower-conductance anion channel (∼25 or ∼50 pS in similar conditions), which only displayed two well-defined substates, at ∼25 and ∼50% of the fully open state. The larger channels were not simple multiples of the lower-conductance channels, but both discriminated poorly, and to a similar extent, between anions and cations (P_Cl ⁻/P_choline ⁺ ∼12, P_Cl ⁻/P_K ⁺∼8). The lower-conductance channel was only minimally selective between different anions (P_{NO 3} ⁻ (1.0)=P_Cl ⁻>P_Br ⁻>P_I ⁻>P_SCN ⁻(0.8)), and its conductance failed to saturate even in high (>1.0 M) activities of KCl. The channels were not obviously voltage dependent, and they were unaffected by 0.5 mM SITS, H₂O₂, propranolol, quinine or amitriptyline, or by 2 mM ATP, or by variations in pH (5.5–8.8). Ca²⁺ and Mg²⁺ did not alter single channel activity, but did modify single current amplitudes in the lower-conductance channel. This effect, together with voltage-dependent substate behavior, is described in the following paper.     

Carbon uptake in Eurasian boreal forests dominates the high-latitude net ecosystem carbon budget

Arctic-boreal landscapes are experiencing profound warming, along with changes in ecosystem moisture status and disturbance from fire. This region is of global importance in terms of carbon feedbacks to climate, yet the sign (sink or source) and magnitude of the Arctic-boreal carbon budget within recent years remains highly uncertain. Here, we provide new estimates of recent (2003–2015) vegetation gross primary productivity (GPP), ecosystem respiration (R_eco), net ecosystem CO₂ exchange (NEE; R_eco − GPP), and terrestrial methane (CH₄) emissions for the Arctic-boreal zone using a satellite data-driven process-model for northern ecosystems (TCFM-Arctic), calibrated and evaluated using measurements from >60 tower eddy covariance (EC) sites. We used TCFM-Arctic to obtain daily 1-km² flux estimates and annual carbon budgets for the pan-Arctic-boreal region. Across the domain, the model indicated an overall average NEE sink of −850 Tg CO₂-C year⁻¹. Eurasian boreal zones, especially those in Siberia, contributed to a majority of the net sink. In contrast, the tundra biome was relatively carbon neutral (ranging from small sink to source). Regional CH₄ emissions from tundra and boreal wetlands (not accounting for aquatic CH₄) were estimated at 35 Tg CH₄-C year⁻¹. Accounting for additional emissions from open water aquatic bodies and from fire, using available estimates from the literature, reduced the total regional NEE sink by 21% and shifted many far northern tundra landscapes, and some boreal forests, to a net carbon source. This assessment, based on in situ observations and models, improves our understanding of the high-latitude carbon status and also indicates a continued need for integrated site-to-regional assessments to monitor the vulnerability of these ecosystems to climate change.     

Structural and serological characterisation of an O-specific polysaccharide from Serratia plymuthica

Abstract The surface polysaccharides of a strain of Serratia plymuthica were characterised and shown to consist of a linear, acidic galactoglucomannan as well as a major and a minor neutral galactan. Immunoblotting results demonstrated cross-reactions between this strain and others with similar galactans ( S. marcescens O16 and O20, Klebsiella O1, and Pasteurella haemolytica T4 and T10).     

Myosin subfragment-1: structure and function of a molecular motor: Current opinion in structural biology 1993, 3:944–952

The ability for directed movement is a fundamental process of all living systems. Molecules designed for such purposes are ubiquitous in eukaryotic cells and have been the focus of intense investigations for many years. Highlighted in this report is the three-dimensional structure of the myosin motor domain—the first such motor protein to be examined by single-crystal X-ray diffraction analysis.     

mtDNA diversity in rhesus monkeys reveals overestimates of divergence time and paraphyly with neighboring species

Reconstructions of the human-African great ape phylogeny by using mitochondrial DNA (mtDNA) have been subject to considerable debate. One confounding factor may be the lack of data on intraspecific variation. To test this hypothesis, we examined the effect of intraspecific mtDNA diversity on the phylogenetic reconstruction of another Plio- Pleistocene radiation of higher primates, the fascicularis group of macaque (Macaca) monkey species. Fifteen endonucleases were used to identify 10 haplotypes of 40-47 restriction sites in M. mulatta, which were compared with similar data for the other members of this species group. Interpopulational, intraspecific mtDNA diversity was large (0.5%- 4.5%), and estimates of divergence time and branching order incorporating this variation were substantially different from those based on single representatives of each species. We conclude that intraspecific mtDNA diversity is substantial in at least some primate species. Consequently, without prior information on the extent of genetic diversity within a particular species, intraspecific variation must be assessed and accounted for when reconstructing primate phylogenies. Further, we question the reliability of hominoid mtDNA phylogenies, based as they are on one or a few representatives of each species, in an already depauperate superfamily of primates.      

Hatching plasticity in a Southeast Asian tree frog mitigates submergence-induced mortality

Environmentally cued hatching has been well-documented in amphibians in response to a wide range of abiotic and biotic factors. The hatching of terrestrial amphibian eggs in response to flooding may be basal within the group, but amphibian lineages in tropical Asia and sub-Saharan Africa have not received as much attention as their Neotropical counterparts. We investigated submergence-induced hatching in Feihyla hansenae, a Rhacophorid tree frog with terrestrial eggs. We quantified natural rates of clutch submergence at our study site in Thailand. Using submergence experiments, we found that embryos are capable of hatching early to escape flooding, and that failure to hatch results in mortality. Among the embryos that were able to hatch early, only the earliest, youngest hatchlings experienced a trade-off in body size that persisted for 6 days, while later, older hatchlings were not significantly smaller than spontaneous hatchlings under control conditions. By incorporating our natural and experimental data into Monte Carlo methods to simulate and compare survival probabilities with and without hatching plasticity, we found an overall 3.1% increase in submergence survival due to hatching plasticity. Our findings support the idea that flooding-induced hatching is widespread across amphibians with terrestrial eggs and highlight the importance of researching understudied tropical regions. As climate change is projected to affect rainfall patterns, the ability of embryos to escape abiotic egg-stage threats may be an indicator of species' ability to flexibly navigate a changing environment.     

Linkage of the Murine Transforming Growth Factor α Gene with Igk, Ly-2, and Fabp1 on Chromosome 6

TGFα is a mitogenic polypeptide found in the conditioned media of transformed cell lines as well as in various solid tumors. Although its physiological role in normal tissues is uncertain, the autocrine action of TGFα on the EGF receptor is postulated to play a role in tumorigenesis. To explore the possibility that pre-existing mouse mutants might have concordance with the mouse TGFα locus (Tgfa) we sought to establish the chromosomal localization of the murine TGFα gene. Using Southern analysis we have detected NcoI and PvuII RFLP in the TGFα gene of progenitor RI mouse strains. These RFLPs have been used to analyze four different RI sets of DNA and to assign Tgfa to the 35-cM region of chromosome 6. Linkage has been established and the data suggest that the distance between Igk and wa-1 anchor loci may be less than 8 cM and that the gene order for the proximal to mid region of mouse chromosome 6 may be: Ggc-Xmmv27-[Brp-1, Lvp-1, Ms6-4]-[Igk, Ly2, Ly3 Odc-rs5, Rn7s-6, Fabp1]-[Tgfa/wa-1]-IL5-R. Homology of synteny has been further defined between the proximal region of mouse chromosome 6 and with the 2p 13-p11 region of human chromosome 2 encompassing TGFA, IGK, CD8A, and FABP1 .     

An optimized live bacterial delivery vehicle safely and efficaciously delivers bacterially transcribed therapeutic nucleic acids

There is an unmet need for delivery platforms that realize the full potential of next-generation nucleic acid therapeutics. The in vivo usefulness of current delivery systems is limited by numerous weaknesses, including poor targeting specificity, inefficient access to target cell cytoplasm, immune activation, off-target effects, small therapeutic windows, limited genetic encoding and cargo capacity, and manufacturing challenges. Here we characterize the safety and efficacy of a delivery platform comprising engineered live, tissue-targeting, non-pathogenic bacteria (Escherichia coli SVC1) for intracellular cargo delivery. SVC1 bacteria are engineered to specifically bind to epithelial cells via a surface-expressed targeting ligand, to allow escape of their cargo from the phagosome, and to have minimal immunogenicity. We describe SVC1's ability to deliver short hairpin RNA (shRNA), localized SVC1 administration to various tissues, and its minimal immunogenicity. To validate the therapeutic potential of SVC1, we used it to deliver influenza-targeting antiviral shRNAs to respiratory tissues in vivo. These data are the first to establish the safety and efficacy of this bacteria-based delivery platform for use in multiple tissue types and as an antiviral in the mammalian respiratory tract. We expect that this optimized delivery platform will enable a variety of advanced therapeutic approaches.