Genome-wide analysis of signatures of selection in populations of African honey bees (Apis mellifera) using new web-based tools

Background

With the development of inexpensive, high-throughput sequencing technologies, it has become feasible to examine questions related to population genetics and molecular evolution of non-model species in their ecological contexts on a genome-wide scale. Here, we employed a newly developed suite of integrated, web-based programs to examine population dynamics and signatures of selection across the genome using several well-established tests, including F_ST, pN/pS, and McDonald-Kreitman. We applied these techniques to study populations of honey bees (Apis mellifera) in East Africa. In Kenya, there are several described A. mellifera subspecies, which are thought to be localized to distinct ecological regions.

Results

We performed whole genome sequencing of 11 worker honey bees from apiaries distributed throughout Kenya and identified 3.6 million putative single-nucleotide polymorphisms. The dense coverage allowed us to apply several computational procedures to study population structure and the evolutionary relationships among the populations, and to detect signs of adaptive evolution across the genome. While there is considerable gene flow among the sampled populations, there are clear distinctions between populations from the northern desert region and those from the temperate, savannah region. We identified several genes showing population genetic patterns consistent with positive selection within African bee populations, and between these populations and European A. mellifera or Asian Apis florea.

Conclusions

These results lay the groundwork for future studies of adaptive ecological evolution in honey bees, and demonstrate the use of new, freely available web-based tools and workflows (http://usegalaxy.org/r/kenyanbee) that can be applied to any model system with genomic information.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1712-0) contains supplementary material, which is available to authorized users.     

Comparison of the Biochemistry and Rates of Synthesis of Mesosomal and Peripheral Membranes in Bacillus subtilis

The membrane vesicle (beaded chain) portion of the mesosomes and peripheral (ghost) membrane of Bacillus subtilis were obtained by protoplast lysis and separated by differential and sucrose gradient centrifugation. Electron microscopy revealed that both fractions were satisfactorily homogeneous. Comparison of the two membrane preparations showed that they were similar with respect to total protein, total phosphorus, and lipid-soluble phosphorus content. Their protein patterns on acrylamide gel electrophreograms did not differ significantly. A possible point of distinction was revealed by a difference spectrum analysis of their cytochromes. The two preparations showed clear quantitative differences in all five of the enzyme activities assayed. Acrylamide gel electrophreograms of peripheral membrane stained for malate dehydrogenase showed four weak isozyme bands, whereas electrophreograms of mesosome membranes exhibited a single strong peak. (A survey of published data on enzymes in mesosome fractions shows a marked lack of correspondence between different species of bacteria.) Comparison of (3)H-acetate incorporation into the two membrane fractions showed that both were labeled at the same rate. Similarly, (35)SO(4) was taken up by both fractions at a comparable rate and was chased from both comparably. Lipid and protein labeling thus indicates that mesosome vesicle membrane is not a precursor or special growing point of peripheral membrane.     

Isolation and partial characterization of single-stranded adenoviral DNA produced during synthesis of adenovirus type 2 DNA.

Single-stranded fragments of adenovirus type 2 DNA were isolated from infected KB cells under conditions which retarded reassociation of complementary sequences but did not denature native viral DNA. Of the total intracellular, virus-specific DNA labeled during a 1-h pulse with tritiated thymidine begining 15 h after infection, about 20% was single stranded when fractionated on hydroxylapatite. This DNA shifted predominantly to the double-stranded fraction on hydroxylapatite during an extended chase incubation, suggesting that it may represent single-stranded DNA in replicating intermediates. Furthermore, the single-stranded DNA annealed nearly equally to both strands of the adenovirus genome. These findings indicate that at least portions of both complementary strands of adenovirus type 2 DNA are exposed as single strands during the period of viral DNA synthesis.     

Affinity labels for the anion-binding site in ovotransferrin

Bromopyruvate, a known alkylating agent, has previously been reported to function as an affinity label for the anion-binding site in the iron-binding protein ovotransferrin [Patch, M.G., & Carrano, C. J. (1982) Biochim. Biophys. Acta 700, 217-220]. However, the present results indicate that hydroxypyruvate also functions in an almost identical manner, which implies that alkylation of a susceptible nucleophile cannot be the mechanism responsible for the covalent attachment of the anion. Model complexes and amino acid analysis of labeled ovotransferrin suggest that initial Schiff base formation, followed by reduction of the imine bond between the affinity anion and a lysine within the locus of the anion-binding site, accounts for the irreversible labeling. As expected, the covalently attached anions render the iron in the ovotransferrin-iron-anion ternary complex much more resistant to loss at low pH. It is proposed that the covalently labeled protein be used to test the hypothesis that iron removal from transferrin occurs by protonation and loss of the anion in low-pH lysosomal vesicles.     

Identification of 2-acylaminothiophene-3-carboxamides as potent inhibitors of FLT3

A series of 2-acylaminothiophene-3-carboxamides has been identified which exhibit potent inhibitory activity against the FLT3 tyrosine kinase. Compound 44 inhibits the isolated enzyme (IC50 = 0.027 microM) and blocks the proliferation of MV4-11 cells (IC50 = 0.41 microM). Structure-activity relationship studies within this series are described in the context of a proposed binding model within the ATP binding site of the enzyme.     

A Second-Site Noncomplementation Screen for Modifiers of Rho1 Signaling during Imaginal Disc Morphogenesis in Drosophila

Background

Rho1 is a small GTPase of the Ras superfamily that serves as the central component in a highly conserved signaling pathway that regulates tissue morphogenesis during development in all animals. Since there is tremendous diversity in the upstream signals that can activate Rho1 as well as the effector molecules that carry out its functions, it is important to define relevant Rho1-interacting genes for each morphogenetic event regulated by this signaling pathway. Previous work from our lab and others has shown that Rho signaling is necessary for the morphogenesis of leg imaginal discs during metamorphosis in Drosophila, although a comprehensive identification of Rho1-interacting genes has not been attempted for this process.

Methodology/Principal Findings

We characterized an amorphic allele of Rho1 that displays a poorly penetrant dominant malformed leg phenotype and is capable of being strongly enhanced by Rho1-interacting heterozygous mutations. We then used this allele in a second-site noncomplementation screen with the Exelixis collection of molecularly defined deficiencies to identify Rho1-interacting genes necessary for leg morphogenesis. In a primary screen of 461 deficiencies collectively uncovering ∼50% of the Drosophila genome, we identified twelve intervals harboring Rho1-interacting genes. Through secondary screening we identified six Rho1-interacting genes including three that were previously identified (RhoGEF2, broad, and stubbloid), thereby validating the screen. In addition, we identified Cdc42, Rheb and Sc2 as novel Rho1-interacting genes involved in adult leg development.

Conclusions/Significance

This screen identified well-known and novel Rho1-interacting genes necessary for leg morphogenesis, thereby increasing our knowledge of this important signaling pathway. We additionally found that Rheb may have a unique function in leg morphogenesis that is independent of its regulation of Tor.