Rapid development of cleaning behaviour by Torresian crows Corvus orru on non-native banteng Bos javanicus in northern Australia

In this paper we report the observation of a rapidly developed vertebrate symbiosis involving ectoparasite cleaning by a native corvid of northern Australia, the Torresian crow Corvus orru , on a recently introduced bovid ungulate, the banteng Bos javanicus . Facultative symbioses benefiting both participants (mutualisms) between birds and mammals appear to be rare, despite the apparent advantages obtained by the participants (ecto-parasite removal from the host and food provision to the cleaner). On three separate dates we observed a total of four C. orru individuals eliciting facilitation behaviours by a total of ten female banteng to assist in the removal of ectoparasites. Our observations document the first-known incidence of facultative cleaning behaviour by a native bird species on a non-native, wild vertebrate that has developed in approximately 150 years since the banteng's introduction to Australia.     

Genome-wide analysis of barcoded Saccharomyces cerevisiae gene-deletion mutants in pooled cultures

The availability of a near-complete (96%) collection of gene-deletion mutants in Saccharomyces cerevisiae greatly facilitates the systematic analyses of gene function in yeast. The unique 20 bp DNA 'barcodes' or 'tags' in each deletion strain enable the individual fitness of thousands of deletion mutants to be resolved from a single pooled culture. Here, we present protocols for the study of pooled cultures of tagged yeast deletion mutants with a tag microarray. This process involves five main steps: pooled growth, isolation of genomic DNA, PCR amplification of the barcodes, array hybridization and data analysis. Pooled deletion screening can be used to study gene function, uncover a compound's mode of action and identify drug targets. In addition to these applications, the general method of studying pooled samples with barcode arrays can also be adapted for use with other types of samples, such as mutant collections in other organisms, short interfering RNA vectors and molecular inversion probes.     

Genetic diversity and population structure identify the potential source of the invasive red clover casebearer moth, <Emphasis Type="Italic">Coleophora deauratella</Emphasis>, in North America

The red clover casebearer, Coleophora deauratella, is an invasive pest of red clover grown for seed in North America. In 2006, an outbreak in Alberta, Canada was discovered that resulted in significant seed losses, while further invasion threatens the world’s largest red clover forage seed production region in Oregon, USA. Prior to the recent outbreak, C. deauratella was thought to be restricted to eastern North America in its invasive range. We sequenced a 615-bp fragment of the mitochondrial cytochrome c oxidase subunit 1 gene, and developed three microsatellite markers to assess the genetic diversity and population structure of C. deauratella in North America and its native range in Europe. We observed signatures of a founder effect in North American populations and a further loss of genetic diversity within Alberta populations. Most genetic differentiation was found between continents, with no evidence of isolation-by-distance within each continent. From the limited number of European populations sampled, a single introduction from Switzerland is the most probable source of North American populations based on similar mitochondrial diversity and lack of population differentiation. Within North America, based on increased genetic diversity compared to the rest of the continent, the first North American record from Ithaca, NY, and the first documented outbreak in southern Ontario in 1989, the initial C. deauratella invasion most likely occurred in southern Ontario, Canada or adjacent states in the USA, followed by transport throughout the continent. This study provides insight into the phylogeographic history of C. deauratella in North America and Europe and may help to identify a regional source of future classical biological control agents.     

Centromere assembly and propagation

Centromere assembly provides a unique example of how elaborate protein structures can be assembled onto DNA, independent of sequence, and then stably propagated through numerous cell divisions. Here, we review the possible epigenetic strategies that organisms ranging from yeast to human use to assemble and propagate active centromeres.     

4.4 Å cryo-EM structure of an enveloped alphavirus Venezuelan equine encephalitis virus

Venezuelan equine encephalitis virus (VEEV), a member of the membrane‐containing Alphavirus genus, is a human and equine pathogen, and has been developed as a biological weapon. Using electron cryo‐microscopy (cryo‐EM), we determined the structure of an attenuated vaccine strain, TC‐83, of VEEV to 4.4 Å resolution. Our density map clearly resolves regions (including E1, E2 transmembrane helices and cytoplasmic tails) that were missing in the crystal structures of domains of alphavirus subunits. These new features are implicated in the fusion, assembly and budding processes of alphaviruses. Furthermore, our map reveals the unexpected E3 protein, which is cleaved and generally thought to be absent in the mature VEEV. Our structural results suggest a mechanism for the initial stage of nucleocapsid core formation, and shed light on the virulence attenuation, host recognition and neutralizing activities of VEEV and other alphavirus pathogens.     

Biochemical and structural studies with prenyl diphosphate analogues provide insights into isoprenoid recognition by protein farnesyl transferase

Protein farnesyl transferase (PFTase) catalyzes the reaction between farnesyl diphosphate and a protein substrate to form a thioether-linked prenylated protein. The fact that many prenylated proteins are involved in signaling processes has generated considerable interest in protein prenyl transferases as possible anticancer targets. While considerable progress has been made in understanding how prenyl transferases distinguish between related target proteins, the rules for isoprenoid discrimination by these enzymes are less well understood. To clarify how PFTase discriminates between FPP and larger prenyl diphosphates, we have examined the interactions between the enzyme and several isoprenoid analogues, GGPP, and the farnesylated peptide product using a combination of biochemical and structural methods. Two photoactive isoprenoid analogues were shown to inhibit yeast PFTase with K(I) values as low as 45 nM. Crystallographic analysis of one of these analogues bound to PFTase reveals that the diphosphate moiety and the two isoprene units bind in the same positions occupied by the corresponding atoms in FPP when bound to PFTase. However, the benzophenone group protrudes into the acceptor protein binding site and prevents the binding of the second (protein) substrate. Crystallographic analysis of geranylgeranyl diphosphate bound to PFTase shows that the terminal two isoprene units and diphosphate group of the molecule map to the corresponding atoms in FPP; however, the first and second isoprene units bulge away from the acceptor protein binding site. Comparison of the GGPP binding mode with the binding of the farnesylated peptide product suggests that the bulkier isoprenoid cannot rearrange to convert to product without unfavorable steric interactions with the acceptor protein. Taken together, these data do not support the "molecular ruler hypotheses". Instead, we propose a "second site exclusion model" in which PFTase binds larger isoprenoids in a fashion that prevents the subsequent productive binding of the acceptor protein or its conversion to product.     

Cabozantinib Inhibits Growth of Androgen-Sensitive and Castration-Resistant Prostate Cancer and Affects Bone Remodeling

Cabozantinib is an inhibitor of multiple receptor tyrosine kinases, including MET and VEGFR2. In a phase II clinical trial in advanced prostate cancer (PCa), cabozantinib treatment improved bone scans in 68% of evaluable patients. Our studies aimed to determine the expression of cabozantinib targets during PCa progression and to evaluate its efficacy in hormone-sensitive and castration-resistant PCa in preclinical models while delineating its effects on tumor and bone. Using immunohistochemistry and tissue microarrays containing normal prostate, primary PCa, and soft tissue and bone metastases, our data show that levels of MET, P-MET, and VEGFR2 are increasing during PCa progression. Our data also show that the expression of cabozantinib targets are particularly pronounced in bone metastases. To evaluate cabozantinib efficacy on PCa growth in the bone environment and in soft tissues we used androgen-sensitive LuCaP 23.1 and castration-resistant C4-2B PCa tumors. In vivo, cabozantinib inhibited the growth of PCa in bone as well as growth of subcutaneous tumors. Furthermore, cabozantinib treatment attenuated the bone response to the tumor and resulted in increased normal bone volume. In summary, the expression pattern of cabozantinib targets in primary and castration-resistant metastatic PCa, and its efficacy in two different models of PCa suggest that this agent has a strong potential for the effective treatment of PCa at different stages of the disease.