Gene Expression and Functional Analyses of Odorant Receptors in Small Hive Beetles (Aethina tumida).

Olfaction is key to many insects. Odorant receptors (ORs) stand among the key chemosensory receptors mediating the detection of pheromones and kairomones. Small hive beetles (SHBs), Aethina tumida, are parasites of social bee colonies and olfactory cues are especially important for host finding. However, how interactions with their hosts may have shaped the evolution of ORs in the SHB remains poorly understood. Here, for the first time, we analyzed the evolution of SHB ORs through phylogenetic and positive selection analyses. We then tested the expression of selected OR genes in antennae, heads, and abdomens in four groups of adult SHBs: colony odor-experienced/-naive males and females. The results show that SHBs experienced both OR gene losses and duplications, thereby providing a first understanding of the evolution of SHB ORs. Additionally, three candidate ORs potentially involved in host finding and/or chemical communication were identified. Significantly different downregulations of ORs between the abdomens of male and female SHBs exposed to colony odors may reflect that these expression patterns might also reflect other internal events, e.g., oviposition. Altogether, these results provide novel insights into the evolution of SHB ORs and provide a valuable resource for analyzing the function of key genes, e.g., for developing biological control. These results will also help in understanding the chemosensory system in SHBs and other beetles

Identification of co-regulated candidate genes by promoter analysis.

EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Uncovering metabolic pathways relevant to phenotypic traits of microbial genomes

A new machine learning-based method is presented here for the identification of metabolic pathways related to specific phenotypes in multiple microbial genomes

Application of a Naïve Bayes Classifier to Assign Polyadenylation Sites from 3\u27 End Deep Sequencing Data: A Dissertation

Cleavage and polyadenylation of a precursor mRNA is important for transcription termination, mRNA stability, and regulation of gene expression. This process is directed by a multitude of protein factors and cis elements in the pre-mRNA sequence surrounding the cleavage and polyadenylation site. Importantly, the location of the cleavage and polyadenylation site helps define the 3’ untranslated region of a transcript, which is important for regulation by microRNAs and RNA binding proteins. Additionally, these sites have generally been poorly annotated. To identify 3’ ends, many techniques utilize an oligo-dT primer to construct deep sequencing libraries. However, this approach can lead to identification of artifactual polyadenylation sites due to internal priming in homopolymeric stretches of adenines. Previously, simple heuristic filters relying on the number of adenines in the genomic sequence downstream of a putative polyadenylation site have been used to remove these sites of internal priming. However, these simple filters may not remove all sites of internal priming and may also exclude true polyadenylation sites. Therefore, I developed a naïve Bayes classifier to identify putative sites from oligo-dT primed 3’ end deep sequencing as true or false/internally primed. Notably, this algorithm uses a combination of sequence elements to distinguish between true and false sites. Finally, the resulting algorithm is highly accurate in multiple model systems and facilitates identification of novel polyadenylation sites

The Energetics of Molecular Adaptation in Transcriptional Regulation

Mutation is a critical mechanism by which evolution explores the functional landscape of proteins. Despite our ability to experimentally inflict mutations at will, it remains difficult to link sequence-level perturbations to systems-level responses. Here, we present a framework centered on measuring changes in the free energy of the system to link individual mutations in an allosteric transcriptional repressor to the parameters which govern its response. We find the energetic effects of the mutations can be categorized into several classes which have characteristic curves as a function of the inducer concentration. We experimentally test these diagnostic predictions using the well-characterized LacI repressor of Escherichia coli, probing several mutations in the DNA binding and inducer binding domains. We find that the change in gene expression due to a point mutation can be captured by modifying only a subset of the model parameters that describe the respective domain of the wild-type protein. These parameters appear to be insulated, with mutations in the DNA binding domain altering only the DNA affinity and those in the inducer binding domain altering only the allosteric parameters. Changing these subsets of parameters tunes the free energy of the system in a way that is concordant with theoretical expectations. Finally, we show that the induction profiles and resulting free energies associated with pairwise double mutants can be predicted with quantitative accuracy given knowledge of the single mutants, providing an avenue for identifying and quantifying epistatic interactions.Comment: 11 pages, 6 figures, supplemental info. available via http://rpgroup.caltech.edu/mwc_mutant

Diversifying selection and functional analysis of interleukin-4 suggests antagonism-driven evolution at receptor-binding interfaces

<p>Abstract</p> <p>Background</p> <p>Interleukin-4 (IL4) is a secreted immunoregulatory cytokine critically involved in host protection from parasitic helminths <abbrgrp><abbr bid="B1">1</abbr></abbrgrp>. Reasoning that helminths may have evolved mechanisms to antagonize IL4 to maximize their dispersal, we explored mammalian IL4 evolution.</p> <p>Results</p> <p>This analysis revealed evidence of diversifying selection at 15 residues, clustered in epitopes responsible for IL4 binding to its Type I and Type II receptors. Such a striking signature of selective pressure suggested either recurrent episodes of pathogen antagonism or ligand/receptor co-evolution. To test the latter possibility, we performed detailed functional analysis of IL4 allotypes expressed by <it>Mus musculus musculus </it>and <it>Mus musculus castaneus</it>, which happen to differ at 5 residues (including three at positively selected sites) in and adjacent to the site 1 epitope that binds the IL4Rα subunit shared by the Type I and Type II IL4 receptors. We show that this intra-species variation affects the ability of IL4 neither to bind IL4 receptor alpha (IL4Rα) nor to signal biological responses through its Type I receptor.</p> <p>Conclusions</p> <p>Our results -- reminiscent of clustered positively selected sites revealing functionally important residues at host-virus interaction interfaces -- are consistent with IL4 having evolved to avoid recurrent pathogen antagonism, while maintaining the capacity to bind and signal through its cognate receptor. This work exposes what may be a general feature of evolutionary conflicts fought by pathogen antagonists at host protein-protein interaction interfaces involved in immune signaling: the emergence of receptor-binding ligand epitopes capable of buffering amino acid variation.</p