Molecular Evolution of Drosophila Cuticular Protein Genes

Several multigene families have been described that together encode scores of structural cuticular proteins in Drosophila, although the functional significance of this diversity remains to be explored. Here I investigate the evolutionary histories of several multigene families (CPR, Tweedle, CPLCG, and CPF/CPFL) that vary in age, size, and sequence complexity, using sequenced Drosophila genomes and mosquito outgroups. My objective is to describe the rates and mechanisms of ‘cuticle-ome’ divergence, in order to identify conserved and rapidly evolving elements. I also investigate potential examples of interlocus gene conversion and concerted evolution within these families during Drosophila evolution. The absolute rate of change in gene number (per million years) is an order of magnitude lower for cuticular protein families within Drosophila than it is among Drosophila and the two mosquito taxa, implying that major transitions in the cuticle proteome have occurred at higher taxonomic levels. Several hotspots of intergenic conversion and/or gene turnover were identified, e.g. some gene pairs have independently undergone intergenic conversion within different lineages. Some gene conversion hotspots were characterized by conversion tracts initiating near nucleotide repeats within coding regions, and similar repeats were found within concertedly evolving cuticular protein genes in Anopheles gambiae. Rates of amino-acid substitution were generally severalfold higher along the branch connecting the Sophophora and Drosophila species groups, and 13 genes have Ka/Ks significantly greater than one along this branch, indicating adaptive divergence. Insect cuticular proteins appear to be a source of adaptive evolution within genera and, at higher taxonomic levels, subject to periods of gene-family expansion and contraction followed by quiescence. However, this relative stasis is belied by hotspots of molecular evolution, particularly concerted evolution, during the diversification of Drosophila. The prominent association between interlocus gene conversion and repeats within the coding sequence of interacting genes suggests that the latter promote strand exchange

Annotation and analysis of a large cuticular protein family with the R&R Consensus in Anopheles gambiae

<p>Abstract</p> <p>Background</p> <p>The most abundant family of insect cuticular proteins, the CPR family, is recognized by the R&R Consensus, a domain of about 64 amino acids that binds to chitin and is present throughout arthropods. Several species have now been shown to have more than 100 CPR genes, inviting speculation as to the functional importance of this large number and diversity.</p> <p>Results</p> <p>We have identified 156 genes in <it>Anopheles gambiae </it>that code for putative cuticular proteins in this CPR family, over 1% of the total number of predicted genes in this species. Annotation was verified using several criteria including identification of TATA boxes, INRs, and DPEs plus support from proteomic and gene expression analyses. Two previously recognized CPR classes, RR-1 and RR-2, form separate, well-supported clades with the exception of a small set of genes with long branches whose relationships are poorly resolved. Several of these outliers have clear orthologs in other species. Although both clades are under purifying selection, the RR-1 variant of the R&R Consensus is evolving at twice the rate of the RR-2 variant and is structurally more labile. In contrast, the regions flanking the R&R Consensus have diversified in amino-acid composition to a much greater extent in RR-2 genes compared with RR-1 genes. Many genes are found in compact tandem arrays that may include similar or dissimilar genes but always include just one of the two classes. Tandem arrays of RR-2 genes frequently contain subsets of genes coding for highly similar proteins (sequence clusters). Properties of the proteins indicated that each cluster may serve a distinct function in the cuticle.</p> <p>Conclusion</p> <p>The complete annotation of this large gene family provides insight on the mechanisms of gene family evolution and clues about the need for so many CPR genes. These data also should assist annotation of other <it>Anopheles </it>genes.</p

The Distribution of GYR- and YLP-Like Motifs in Drosophila Suggests a General Role in Cuticle Assembly and Other Protein-Protein Interactions

Background: Arthropod cuticle is composed predominantly of a self-assembling matrix of chitin and protein. Genes encoding structural cuticular proteins are remarkably abundant in arthropod genomes, yet there has been no systematic survey of conserved motifs across cuticular protein families. Methodology/Principal Findings: Two short sequence motifs with conserved tyrosines were identified in Drosophila cuticular proteins that were similar to the GYR and YLP Interpro domains. These motifs were found in members of the CPR, Tweedle, CPF/CPFL, and (in Anopheles gambiae) CPLCG cuticular protein families, and the Dusky/Miniature family of cuticleassociated proteins. Tweedle proteins have a characteristic motif architecture that is shared with the Drosophila protein GCR1 and its orthologs in other species, suggesting that GCR1 is also cuticular. A resilin repeat, which has been shown to confer elasticity, matched one of the motifs; a number of other Drosophila proteins of unknown function exhibit a motif architecture similar to that of resilin. The motifs were also present in some proteins of the peritrophic matrix and the eggshell, suggesting molecular convergence among distinct extracellular matrices. More surprisingly, gene regulation, development, and proteolysis were statistically over-represented ontology terms for all non-cuticular matches in Drosophila. Searches against other arthropod genomes indicate that the motifs are taxonomically widespread. Conclusions: This survey suggests a more general definition for GYR and YLP motifs and reveals their contribution to severa

Complementary and Alternative Medicine (CAM) Attitudes and Competencies of Nursing Students and Faculty: Results of Integrating CAM Into the Nursing Curriculum

As part of the National Center for Complementary and Alternative Medicine (CAM) R25 Education Grant Program, a faculty development program for integrating CAM into the nursing curriculum was instituted in 2003-2006. The Integrating CAM program comprised a number of elements; the primary strategy included a series of 4-week didactic and experiential summer CAM “camps,” attended by a total of 27 faculty members. Camps were designed to influence faculty integration of CAM material into course offerings. The Integrating CAM program was evaluated via a series of faculty and student surveys regarding CAM competencies, attitudes, and perceptions. For more than half of the faculty (out of the 43 who responded), the program yielded a moderate-to-strong influence on incorporation of CAM material into course content; and moderate-to-great increases in both enthusiasm for CAM and perceived CAM knowledge gains. Students at all levels (undergraduate, masters, doctoral; n = 184) reported that their courses contained CAM content; for 70% of students, their CAM knowledge increased; for 50% of students, level of CAM interest increased. Self-reported student CAM competencies were significantly greater in 2006-2007 (n = 191) than in 2003-2004 (n = 143). Results support the strategy of broadly infusing the nursing curriculum with CAM content via faculty development

Genomic survey of the ectoparasitic mite Varroa destructor, a major pest of the honey bee Apis mellifera

Background: The ectoparasitic mite Varroa destructor has emerged as the primary pest of domestic honey bees (Apis mellifera). Here we present an initial survey of the V. destructor genome carried out to advance our understanding of Varroa biology and to identify new avenues for mite control. This sequence survey provides immediate resources for molecular and population-genetic analyses of Varroa-Apis interactions and defines the challenges ahead for a comprehensive Varroa genome project. Results: The genome size was estimated by flow cytometry to be 565 Mbp, larger than most sequenced insects but modest relative to some other Acari. Genomic DNA pooled from similar to 1,000 mites was sequenced to 4.3x coverage with 454 pyrosequencing. The 2.4 Gbp of sequencing reads were assembled into 184,094 contigs with an N50 of 2,262 bp, totaling 294 Mbp of sequence after filtering. Genic sequences with homology to other eukaryotic genomes were identified on 13,031 of these contigs, totaling 31.3 Mbp. Alignment of protein sequence blocks conserved among V. destructor and four other arthropod genomes indicated a higher level of sequence divergence within this mite lineage relative to the tick Ixodes scapularis. A number of microbes potentially associated with V. destructor were identified in the sequence survey, including similar to 300 Kbp of sequence deriving from one or more bacterial species of the Actinomycetales. The presence of this bacterium was confirmed in individual mites by PCR assay, but varied significantly by age and sex of mites. Fragments of a novel virus related to the Baculoviridae were also identified in the survey. The rate of single nucleotide polymorphisms (SNPs) in the pooled mites was estimated to be 6.2 x 10(-5)per bp, a low rate consistent with the historical demography and life history of the species. Conclusions: This survey has provided general tools for the research community and novel directions for investigating the biology and control of Varroa mites. Ongoing development of Varroa genomic resources will be a boon for comparative genomics of under-represented arthropods, and will further enhance the honey bee and its associated pathogens as a model system for studying host-pathogen interactions

Robot Learning through Crowd-Based Games

The field of robot learning from demonstration focuses on algorithms that enable a robot to learn new abilities from examples provided by a human teacher. We develop a system that enables users to remotely control a KUKA youBot robot to play a game of Whack-A-Mole through a common web browser. We collect data on how users play and utilize a decision tree learning algorithm to teach the robot to play autonomously. We find that filtering data by score does not significantly improve learned robot performance over using all data. Using A/B testing techniques, we find that motivational game features noticeably improve the quantity and quality of collected data

Summary-figure-of-nucleotide-distance-conserved-regions

This figure summarizes the nucleotide distances from the spreadsheet "conserved-primer-candidates-for-orthologous-segments.xls" in order to better guide the selection of loci for molecular phylogenetics. Loci with greater nucleotide distances may be more informative for analysis of closely related species

Conserved signal peptides of AMPs

Sequence logos of the highly conserved signal peptide and acidic propiece region for each species, based on aligned cluster sequences. Standard amino-acid symbols are used, with dark font representing more acidic residues. The predicted cleavage site is C-terminal of the conserved cysteine residue. Position 24 of the P. regilla alignment is blank because the majority of sequences had a gap at this alignment position

antimicrobial-peptide-clusters

A FASTA-formatted file containing aligned representative sequences for each cluster of antimicrobial peptide reads and/or contigs. The sequence headers correspond to the phylogeny in Figure 1