Production of aggressive electrocommunication signals to progressively realistic social stimuli in male Apteronotus leptorhynchus

Brown ghost knife fish, Apteronotus leptorhynchus, produce a continuous electric organ discharge (EOD) that they use for communication. While interacting aggressively, males also emit brief EOD modulations termed chirps. The simplicity of this behaior and its underlying neural circuitry has made it an important model system in neuroethology. Chirping is typically assayed by confining a fish in a tube (‘chirp chamber’) and presenting it with sine wave electrical stimuli that partially mimic EODs of other fish. We presented male fish with progressively more realistic social stimuli to examine whether some of the stimulus complexities during dyadic interaction influence the production of chirps. In a chirp chamber, fish chirped less to a recording of an EOD containing chirps than to a recording of an EOD alone and to sine wave stimuli. Free‐swimming fish chirped more to stimulus fish than to sine wave stimuli presented through electrodes. Fish chirped more when interacting directly than when interacting across a perforated barrier. Together, these studies demonstrate that the presence of chirps, electric field complexity, and/or non‐electric social stimuli are important in eliciting chirp production in brown ghosts.     

A new basal sauropodiform from South Africa and the phylogenetic relationships of basal sauropodomorphs

We present a new medium‐sized basal sauropodomorph, S efapanosaurus zastronensis gen. et sp. nov. , from the Upper Triassic?Lower Jurassic Elliot Formation of South Africa. It is represented by parts of the postcranial skeleton of at least four individuals, including: cervical, dorsal, sacral and caudal vertebrae, most of the forelimb, and part of the hindlimb. Sefapanosaurus bears several autapomorphies of the astragalus, and referred material also shows autapomorphic features. The inclusion of Sefapanosaurus in a phylogenetic analysis places it within the group of sauropodomorphs more closely related to sauropods than to Massospondylus (i.e. Sauropodiformes), increasing the currently known diversity of the so‐called ‘transitional forms’ leading to Sauropoda. Character optimization revealed the presence of several features that are common for taxa placed within the transitional branches basal to Sauropoda. Sefapanosaurus, together with other transitional sauropodomorphs reported during the last decade, highlights the importance of Gondwanan taxa for understanding the palaeobiodiversity, global distribution, and macroevolutionary changes in the group related to the rise of sauropods. © 2015 The Linnean Society of London     

Crystal Structures of Ricin Toxin's Enzymatic Subunit (RTA) in Complex with Neutralizing and Non-Neutralizing Single-Chain Antibodies

Ricin is a select agent toxin and a member of the RNA N-glycosidase family of medically important plant and bacterial ribosome-inactivating proteins. In this study, we determined X-ray crystal structures of the enzymatic subunit of ricin (RTA) in complex with the antigen binding domains (VHH) of five unique single-chain monoclonal antibodies that differ in their respective toxin-neutralizing activities. None of the VHHs made direct contact with residues involved in RTA's RNA N-glycosidase activity or induced notable allosteric changes in the toxin's subunit. Rather, the five VHHs had overlapping structural epitopes on the surface of the toxin and differed in the degree to which they made contact with prominent structural elements in two folding domains of the RTA. In general, RTA interactions were influenced most by the VHH CDR3 (CDR, complementarity-determining region) elements, with the most potent neutralizing antibody having the shortest and most conformationally constrained CDR3. These structures provide unique insights into the mechanisms underlying toxin neutralization and provide critically important information required for the rational design of ricin toxin subunit vaccines.     

Simple Methods for Generating and Detecting Locus-Specific Mutations Induced with TALENs in the Zebrafish Genome

The zebrafish is a powerful experimental system for uncovering gene function in vertebrate organisms. Nevertheless, studies in the zebrafish have been limited by the approaches available for eliminating gene function. Here we present simple and efficient methods for inducing, detecting, and recovering mutations at virtually any locus in the zebrafish. Briefly, double-strand DNA breaks are induced at a locus of interest by synthetic nucleases, called TALENs. Subsequent host repair of the DNA lesions leads to the generation of insertion and deletion mutations at the targeted locus. To detect the induced DNA sequence alterations at targeted loci, genomes are examined using High Resolution Melt Analysis, an efficient and sensitive method for detecting the presence of newly arising sequence polymorphisms. As the DNA binding specificity of a TALEN is determined by a custom designed array of DNA recognition modules, each of which interacts with a single target nucleotide, TALENs with very high target sequence specificities can be easily generated. Using freely accessible reagents and Web-based software, and a very simple cloning strategy, a TALEN that uniquely recognizes a specific pre-determined locus in the zebrafish genome can be generated within days. Here we develop and test the activity of four TALENs directed at different target genes. Using the experimental approach described here, every embryo injected with RNA encoding a TALEN will acquire targeted mutations. Multiple independently arising mutations are produced in each growing embryo, and up to 50% of the host genomes may acquire a targeted mutation. Upon reaching adulthood, approximately 90% of these animals transmit targeted mutations to their progeny. Results presented here indicate the TALENs are highly sequence-specific and produce minimal off-target effects. In all, it takes about two weeks to create a target-specific TALEN and generate growing embryos that harbor an array of germ line mutations at a pre-specified locus.     

Improved delivery of broadly neutralizing antibodies by nanocapsules suppresses SHIV infection in the CNS of infant rhesus macaques

Broadly neutralizing antibodies (bNAbs) directed to HIV-1 have shown promise at suppressing viremia in animal models. However, the use of bNAbs for the central nervous system (CNS) infection is confounded by poor penetration of the blood brain barrier (BBB). Typically, antibody concentrations in the CNS are extremely low; with levels in cerebrospinal fluid (CSF) only 0.1% of blood concentrations. Using a novel nanotechnology platform, which we term nanocapsules, we show effective transportation of the human bNAb PGT121 across the BBB in infant rhesus macaques upon systemic administration up to 1.6% of plasma concentration. We demonstrate that a single dose of PGT121 encased in nanocapsules when delivered at 48h post-infection delays early acute infection with SHIV_SF162P3 in infants, with one of four animals demonstrating viral clearance. Importantly, the nanocapsule delivery of PGT121 improves suppression of SHIV infection in the CNS relative to controls.     

ErbB2 directly activates the exchange factor Dock7 to promote Schwann cell migration

The cellular events that precede myelination in the peripheral nervous system require rapid and dynamic morphological changes in the Schwann cell. These events are thought to be mainly controlled by axonal signals. But how signals on the axons are coordinately organized and transduced to promote proliferation, migration, radial sorting, and myelination is unknown. We describe that the axonal signal neuregulin-1 (NRG1) controls Schwann cell migration via activation of the atypical Dock180-related guanine nucleotide exchange factor (GEF) Dock7 and subsequent activation of the Rho guanine triphosphatases (GTPases) Rac1 and Cdc42 and the downstream c-Jun N-terminal kinase. We show that the NRG1 receptor ErbB2 directly binds and activates Dock7 by phosphorylating Tyr-1118. Dock7 knockdown, or expression of Dock7 harboring the Tyr-1118-to-Phe mutation in Schwann cells, attenuates the effects of NRG1. Thus, Dock7 functions as an intracellular substrate for ErbB2 to promote Schwann cell migration. This provides an unanticipated mechanism through which ligand-dependent tyrosine phosphorylation can trigger the activation of Rho GTPase-GEFs of the Dock180 family.     

Ecological Correlates to Lemur Community Structure in Southeast Madagascar

The Fandriana-Marolambo forest corridor is one of the largest (ca. 250,000 ha) and least explored tracts of unprotected forest in southeast Madagascar. Although published range maps show continuous distributions for many lemurs throughout the region, there are few data on lemur community structure in the corridor. We aimed to determine lemur community structure, with its ecological correlates (altitude, agriculture, selective logging, and hunting), in the Fandriana-Marolambo forest corridor. We surveyed 7 sites and sighted 4 nocturnal taxa (Avahi laniger, Cheirogaleus major, Lepilemur mustelinus, and Microcebus rufus) and 6 diurnal taxa (Eulemur rubriventer, E. fulvus rufus, E. f. fulvus, Propithecus diadema edwardsi, Hapalemur griseus griseus, and Varecia variegata variegata). Composition of the lemur community was broadly similar to that of nearby protected areas (Ranomafana and Mantadia National Parks). However, we sighted no Hapalemur aureus, H. simus, or Indri indri, and observed Propithecus diadema edwardsi and Varecia variegata variegata at only 1 site each. We sighted an apparent hybrid form of Eulemur fulvus fulvus and E. f. rufus that may represent a new hybrid zone for lemurs. After testing for spatial autocorrelation, lemur diversity correlates negatively with altitude and agricultural intensity. Though the Government of Madagascar is assessing the corridor as a new national park, we suggest conservation plans for local lemurs are complicated by population isolation and lack of data on minimum viable size of the proposed protected area.     

Selective measurement of NAPE-PLD activity via a PLA1/2-resistant fluorogenic N-acyl-phosphatidylethanolamine analog

N-acyl-phosphatidylethanolamine (NAPE)-hydrolyzing phospholipase D (NAPE-PLD) is a zinc metallohydrolase enzyme that converts NAPEs to bioactive N-acyl-ethanolamides. Altered NAPE-PLD activity may contribute to pathogenesis of obesity, diabetes, atherosclerosis, and neurological diseases. Selective measurement of NAPE-PLD activity is challenging, however, because of alternative phospholipase pathways for NAPE hydrolysis. Previous methods to measure NAPE-PLD activity involved addition of exogenous NAPE followed by TLC or LC/MS/MS, which are time and resource intensive. Recently, NAPE-PLD activity in cells has been assayed using the fluorogenic NAPE analogs PED-A1 and PED6, but these substrates also detect the activity of serine hydrolase-type lipases PLA₁ and PLA₂. To create a fluorescence assay that selectively measured cellular NAPE-PLD activity, we synthesized an analog of PED-A1 (flame-NAPE) where the sn-1 ester bond was replaced with an N-methyl amide to create resistance to PLA₁ hydrolysis. Recombinant NAPE-PLD produced fluorescence when incubated with either PED-A1 or flame-NAPE, whereas PLA₁ only produced fluorescence when incubated with PED-A1. Furthermore, fluorescence in HepG2 cells using PED-A1 could be partially blocked by either biothionol (a selective NAPE-PLD inhibitor) or tetrahydrolipstatin (an inhibitor of a broad spectrum of serine hydrolase-type lipases). In contrast, fluorescence assayed in HepG2 cells using flame-NAPE could only be blocked by biothionol. In multiple cell types, the phospholipase activity detected using flame-NAPE was significantly more sensitive to biothionol inhibition than that detected using PED-A1. Thus, using flame-NAPE to measure phospholipase activity provides a rapid and selective method to measure NAPE-PLD activity in cells and tissues.     

Questioning Ramayanas: A South Asian Tradition/Seeking Mahadevi: Constructing the Identities of the Hindu Great Goddess.

Questioning Ramayanas:. South Asian Tradition. Paula Richman. ed. Berkeley: University of California Press, 2001. 452 pp.
Seeking Mahadevi: Constructing the Identities of the Hindu Great Goddess. Tracy Pintchman. ed. Albany: State University of New York Press, 2001. 254 pp.