Successful artificial insemination in the corn snake,Elaphe gutatta,using fresh and cooled semen

The design and implementation of assisted reproductive technology to improve genetic diversity and augment captive populations is an important but rarely applied research field in reptiles. Using the corn snake (Elaphe gutatta) as a model, the Henry Doorly Zoo recently produced offspring born as a result of artificial insemination using both fresh, diluted semen, and diluted semen stored at refrigeration for 3 days. Semen was collected noninvasively from sexually mature male corn snakes using a gentle massaging technique, extended in medium then inseminated into the oviducts of adult females. Using molecular genetic techniques to confirm or refute the success of the insemination using primers developed for the black rat snake, Elaphe obsolete, all possible parents and offspring genotypes were evaluated. A paternity‐by‐exclusion analysis verified that the offspring were in fact a result of artificial insemination. Zoo Biol 26:363–369, 2007. © 2007 Wiley‐Liss, Inc.     

Biophysical characterization of an integrin-targeted lipopolyplex gene delivery vector

Nonviral gene delivery vectors now show good therapeutic potential: however, detailed characterization of the composition and macromolecular organization of such particles remains a challenge. This paper describes experiments to elucidate the structure of a ternary, targeted, lipopolyplex synthetic vector, the LID complex. This consists of a lipid component, Lipofectin (L) (1:1 DOTMA:DOPE), plasmid DNA (D), and a dual-function, cationic peptide component (I) containing DNA condensation and integrin-targeting sequences. Fluorophore-labeled lipid, peptide, and DNA components were used to formulate the vector, and the stoichiometry of the particles was established by fluorescence correlation spectroscopy (FCS). The size of the complex was measured by FCS, and the sizes of LID, L, LD, and ID complexes were measured by dynamic light scattering (DLS). Fluorescence quenching experiments and freeze-fracture electron microscopy were then used to demonstrate the arrangement of the lipid, peptide, and DNA components within the complex. These experiments showed that the cationic portion of the peptide, I, interacts with the plasmid DNA, resulting in a tightly condensed DNA-peptide inner core; this is surrounded by a disordered lipid layer, from which the integrin-targeting sequence of the peptide partially protrudes.     

A New, Commercially Valuable Chanterelle Species, Cantharellus californicus sp. nov., Associated with Live Oak in California, USA

A New, Commercially Valuable Chanterelle Species, Cantharellus californicus sp. nov., Associated with Live Oak in California, USA. The prominent golden chanterelle of California’s oak woodlands is characterized as a new species, Cantharellus californicus sp. nov., using molecular and morphological data. Our observations indicate that it is the largest Cantharellus species in the world, with individual sporocraps commonly weighing 1/2 kilogram (kg) (or 1 pound) or more when mature. Other Cantharellus species in California are compared and evaluated, including their known ectomycorrhizal hosts. The California oak chanterelle is an economically valuable species, and some observations on its commercial harvest are presented.     

Fragment-based design,synthesis, biological evaluation,and SAR of 1H-benzo[d]imidazol-2-yl)-1H-indazol derivatives as potent PDK1 inhibitors

In this work, we describe the use of the rule of 3 fragment-based strategies from biochemical screening data of 1100 in-house, small, low molecular weight fragments. The sequential combination of in silico fragment hopping and fragment linking based on S160/Y161/A162 hinge residues hydrogen bonding interactions leads to the identification of novel 1H-benzo[d]imidazol-2-yl)-1H-indazol class of Phosphoinositide-Dependent Kinase-1 (PDK1) inhibitors. Consequent SAR and follow-up screening data led to the discovery of two potent PDK1 inhibitors: compound 32 and 35, with an IC₅₀ of 80?nM and 94?nM, respectively. Further biological evaluation showed that, at the low nanomolar concentration, the drug had potent ability to inhibit phosphorylation of AKT and p70S6, and selectively kill the cancer cells with mutations in both PTEN and PI3K. The microarray data showed that DUSP6, DUSP4, and FOSL1 were down-regulated in the sensitive cell lines with the compound treatment. The in vivo test showed that 35 can significantly inhibit tumor growth without influencing body weight growth. Our results suggest that these compounds, especially 35, merit further pre-clinical evaluation.     

Disrupting Circadian Homeostasis of Sympathetic Signaling Promotes Tumor Development in Mice

Background

Cell proliferation in all rapidly renewing mammalian tissues follows a circadian rhythm that is often disrupted in advanced-stage tumors. Epidemiologic studies have revealed a clear link between disruption of circadian rhythms and cancer development in humans. Mice lacking the circadian genes Period1 and 2 (Per) or Cryptochrome1 and 2 (Cry) are deficient in cell cycle regulation and Per2 mutant mice are cancer-prone. However, it remains unclear how circadian rhythm in cell proliferation is generated in vivo and why disruption of circadian rhythm may lead to tumorigenesis.

Methodology/Principal Findings

Mice lacking Per1 and 2, Cry1 and 2, or one copy of Bmal1, all show increased spontaneous and radiation-induced tumor development. The neoplastic growth of Per-mutant somatic cells is not controlled cell-autonomously but is dependent upon extracellular mitogenic signals. Among the circadian output pathways, the rhythmic sympathetic signaling plays a key role in the central-peripheral timing mechanism that simultaneously activates the cell cycle clock via AP1-controlled Myc induction and p53 via peripheral clock-controlled ATM activation. Jet-lag promptly desynchronizes the central clock-SNS-peripheral clock axis, abolishes the peripheral clock-dependent ATM activation, and activates myc oncogenic potential, leading to tumor development in the same organ systems in wild-type and circadian gene-mutant mice.

Conclusions/Significance

Tumor suppression in vivo is a clock-controlled physiological function. The central circadian clock paces extracellular mitogenic signals that drive peripheral clock-controlled expression of key cell cycle and tumor suppressor genes to generate a circadian rhythm in cell proliferation. Frequent disruption of circadian rhythm is an important tumor promoting factor.     

Preparation of 14C-Labeled Sterigmatocystin in Liquid Media

¹⁴C-labeled sterigmatocystin was prepared from surface cultures of Aspergillus versicolor A-18074 maintained in liquid media by multiple additions of [1-¹⁴C]acetate to the cultures. The highest yield of 7.75 mg/10 ml was found with a sucrose-asparagine-ammonium medium in which more than 3% of the radioactivity of the added [1-¹⁴C]acetate was recovered in the purified [ring-¹⁴C] sterigmatocystin. The method offers an easy way to prepare ¹⁴C-labeled sterigmatocystin for studies of this mycotoxin.     

On being a Gulf veteran: an anthropological perspective

There is no doubt that Gulf service has affected the well-being of some of the members of the UK armed forces who served in that conflict, yet the reason for this remain unclear. At present, the debate surrounding Gulf War Syndrome (GWS) has become stagnant and highly polarized. This paper argues that a new perspective is needed to further improve our understanding of the problem and suggests that the methods and theories of anthropology, with its focus on nuances and subtleties, can provide new insights. Data were generated from 14 months of ethnographic fieldwork in the UK including participant observation, semi-structured interviews and document analysis. Anthropology provides a unique way of approaching and understanding somatic symptoms and suggests that GWS symptom reporting can be seen as a form of communication. The work focuses on the sufferers' accounts, the symptoms themselves and the context within which we find them in order to better understand what was being expressed and commented upon. Although necessary to contextualize GWS through situating it among other emergent illnesses and widespread health beliefs, this paper shows there is a need to bring back the particular. This work seeks to make sense of the cultural circumstances, specific and general, which gave rise to the illness.     

Loss of Apm1, the micro1 subunit of the clathrin-associated adaptor-protein-1 complex, causes distinct phenotypes and synthetic lethality with calcineurin deletion in fission yeast

Calcineurin is a highly conserved regulator of Ca(2+) signaling in eukaryotes. In fission yeast, calcineurin is not essential for viability but is required for cytokinesis and Cl(-) homeostasis. In a genetic screen for mutations that are synthetically lethal with calcineurin deletion, we isolated a mutant, cis1-1/apm1-1, an allele of the apm1(+) gene that encodes a homolog of the mammalian micro1A subunit of the clathrin-associated adaptor protein-1 (AP-1) complex. The cis1-1/apm1-1 mutant as well as the apm1-deleted (Deltaapm1) cells showed distinct phenotypes: temperature sensitivity; tacrolimus (FK506) sensitivity; and pleiotropic defects in cytokinesis, cell integrity, and vacuole fusion. Electron micrographs revealed that Deltaapm1 cells showed large vesicular structures associated with Golgi stacks and accumulated post-Golgi secretory vesicles. Deltaapm1 cells also showed the massive accumulation of the exocytic v-SNARE Syb1 in the Golgi/endosomes and a reduced secretion of acid phosphatase. These phenotypes observed in apm1 mutations were accentuated upon temperature up-shift and FK506 treatment. Notably, Apm1-GFP localized to the Golgi/endosomes, the spindle pole bodies, and the medial region. These findings suggest a role for Apm1 associated with the Golgi/endosome function, thereby affecting various cellular processes, including secretion, cytokinesis, vacuole fusion, and cell integrity and also suggest that calcineurin is involved in these events.