Expression Profiles Reveal Parallel Evolution of Epistatic Interactions Involving the CRP Regulon in Escherichia coli

The extent and nature of epistatic interactions between mutations are issues of fundamental importance in evolutionary biology. However, they are difficult to study and their influence on adaptation remains poorly understood. Here, we use a systems-level approach to examine epistatic interactions that arose during the evolution of Escherichia coli in a defined environment. We used expression arrays to compare the effect on global patterns of gene expression of deleting a central regulatory gene, crp. Effects were measured in two lineages that had independently evolved for 20,000 generations and in their common ancestor. We found that deleting crp had a much more dramatic effect on the expression profile of the two evolved lines than on the ancestor. Because the sequence of the crp gene was unchanged during evolution, these differences indicate epistatic interactions between crp and mutations at other loci that accumulated during evolution. Moreover, a striking degree of parallelism was observed between the two independently evolved lines; 115 genes that were not crp-dependent in the ancestor became dependent on crp in both evolved lines. An analysis of changes in crp dependence of well-characterized regulons identified a number of regulatory genes as candidates for harboring beneficial mutations that could account for these parallel expression changes. Mutations within three of these genes have previously been found and shown to contribute to fitness. Overall, these findings indicate that epistasis has been important in the adaptive evolution of these lines, and they provide new insight into the types of genetic changes through which epistasis can evolve. More generally, we demonstrate that expression profiles can be profitably used to investigate epistatic interactions

Evolution of Mutational Robustness in an RNA Virus

Mutational (genetic) robustness is phenotypic constancy in the face of mutational changes to the genome. Robustness is critical to the understanding of evolution because phenotypically expressed genetic variation is the fuel of natural selection. Nonetheless, the evidence for adaptive evolution of mutational robustness in biological populations is controversial. Robustness should be selectively favored when mutation rates are high, a common feature of RNA viruses. However, selection for robustness may be relaxed under virus co-infection because complementation between virus genotypes can buffer mutational effects. We therefore hypothesized that selection for genetic robustness in viruses will be weakened with increasing frequency of co-infection. To test this idea, we used populations of RNA phage φ6 that were experimentally evolved at low and high levels of co-infection and subjected lineages of these viruses to mutation accumulation through population bottlenecking. The data demonstrate that viruses evolved under high co-infection show relatively greater mean magnitude and variance in the fitness changes generated by addition of random mutations, confirming our hypothesis that they experience weakened selection for robustness. Our study further suggests that co-infection of host cells may be advantageous to RNA viruses only in the short term. In addition, we observed higher mutation frequencies in the more robust viruses, indicating that evolution of robustness might foster less-accurate genome replication in RNA viruses

Evolution of Highly Polymorphic T Cell Populations in Siblings with the Wiskott-Aldrich Syndrome

Population level evolutionary processes can occur within a single organism when the germ line contains a mutation that confers a cost at the level of the cell. Here we describe how multiple compensatory mutations arose through a within-individual evolutionary process in two brothers with the immune deficiency Wiskott-Aldrich Syndrome (WAS). As a result, both brothers have T lymphocyte populations that are highly polymorphic at the locus of the germ line defect, and no single allele achieves fixation. WASP, the gene product affected in this disease, is specific to white blood cells where it is responsible for regulating actin cytoskeleton dynamics in a wide range of cellular responses. The brothers inherited a rare allele predicted to result in truncated WASP lacking the carboxy-terminal VCA domains, the region that directly catalyzes actin filament generation. Although the brothers' T cell populations are highly polymorphic, all share a corrective effect relative to the inherited allele in that they restore the VCA domain. This indicates massive selection against the truncated germ line allele. No single somatic allele becomes fixed in the circulating T cell population of either brother, indicating that a regulated step in maturation of the affected cell lineage is severely compromised by the germ line allele. Based on the finding of multiple somatic mutations, the known maturation pathway for T-lineage cells and the known defects of T cells and precursor thymocytes in mice with truncated WASP, we hypothesize that the presence of truncated WASP (WASPΔVCA) confers an extreme disadvantage in early developing thymocytes, above and beyond the known cost of absence of full-length WASP, and that the disadvantage likely occurs through dominant negative competition of WASPΔVCA with N-WASP, a protein that otherwise partially compensates for WASP absence in developing thymocytes

Treating cofactors can reverse the expansion of a primary disease epidemic

<p>Abstract</p> <p>Background</p> <p>Cofactors, "nuisance" conditions or pathogens that affect the spread of a primary disease, are likely to be the norm rather than the exception in disease dynamics. Here we present a "simplest possible" demographic model that incorporates two distinct effects of cofactors: that on the transmission of the primary disease from an infected host bearing the cofactor, and that on the acquisition of the primary disease by an individual that is not infected with the primary disease but carries the cofactor.</p> <p>Methods</p> <p>We constructed and analyzed a four-patch compartment model that accommodates a cofactor. We applied the model to HIV spread in the presence of the causal agent of genital schistosomiasis, <it>Schistosoma hematobium</it>, a pathogen commonly co-occurring with HIV in sub-Saharan Africa.</p> <p>Results</p> <p>We found that cofactors can have a range of effects on primary disease dynamics, including shifting the primary disease from non-endemic to endemic, increasing the prevalence of the primary disease, and reversing demographic growth when the host population bears only the primary disease to demographic decline. We show that under parameter values based on the biology of the HIV/<it>S. haematobium </it>system, reduction of the schistosome-bearing subpopulations (e.g. through periodic use of antihelminths) can slow and even reverse the spread of HIV through the host population.</p> <p>Conclusions</p> <p>Typical single-disease models provide estimates of future conditions and guidance for direct intervention efforts relating only to the modeled primary disease. Our results suggest that, in circumstances under which a cofactor affects the disease dynamics, the most effective intervention effort might not be one focused on direct treatment of the primary disease alone. The cofactor model presented here can be used to estimate the impact of the cofactor in a particular disease/cofactor system without requiring the development of a more complicated model which incorporates many other specific aspects of the chosen disease/cofactor pair. Simulation results for the HIV/<it>S. haematobium </it>system have profound implications for disease management in developing areas, in that they provide evidence that in some cases treating cofactors may be the most successful and cost-effective way to slow the spread of primary diseases.</p

Data from: Life in the cystic fibrosis upper respiratory tract influences competitive ability of the opportunistic pathogen Pseudomonas aeruginosa

Understanding characteristic differences between host-associated and free-living opportunistic pathogens can provide insight into the fundamental requirements for success after dispersal to the host environment, and more generally into the ecological and evolutionary processes by which populations respond to simultaneous selection on complex interacting traits. We examined how cystic fibrosis (CF) associated and environmental isolates of the opportunistic pathogen Pseudomonas aeruginosa differ in the production of an ecologically important class of proteinaceous toxins known as bacteriocins, and how overall competitive ability depends on the production of and resistance to these bacteriocins. We determined bacteriocin gene content in a diverse collection of environmental and CF isolates and measured bacteriocin-mediated inhibition, resistance, and the outcome of competition in a shared environment between all possible pairs of these isolates at 25 °C and 37 °C. Although CF isolates encoded significantly more bacteriocin genes, our phenotypic assays suggest that they have diminished bacteriocin-mediated killing and resistance capabilities relative to environmental isolates, regardless of incubation temperature. Notably, however, although bacteriocin killing and resistance profiles significantly predicted head to head competitive outcomes, CF and environmental isolates did not differ significantly in their competitive ability. This suggests that the contribution of bacteriocins to competitive ability involves selection on other traits that may be pleiotropically linked to interference competition mediated by bacteriocins

Fourteen Somatic Patient Alleles Identified by Cloning.

<p>Top: Alleles 1–7. Shown is the affected segment of exon 10 of normal <i>WASP</i> (N), the patients' germ line allele (G) and somatic alleles #1–7 (identified on the right). The arrow marks the germ line 1305G deletion. These 7 alleles encode WASP with deletions in the linker region that minimally impinge on the upstream proline-rich region (orange) and do not impinge on the downstream verprolin homology (V) domain (green). Features favorable to mutations include the high GC content and particularly nucleotides 1299–1316 (yellow), which may form a hairpin loop stabilized by five G-C bonds, and tandem 4 nucleotide repeats (underlined) that could predispose to deletion by slipped strand mispairing. Bottom: Alleles # 8–14. Shown is the entire exon 10 with 1305G marked by an arrow. Patient allele #3 is repeated to facilitate comparison. Deleted segments are shown as black, out-of-frame segments as crosshatched. Numbering is relative to normal <i>WASP</i>. Further details are in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0003444#pone-0003444-t002" target="_blank">Table 2</a>.</p

WASP in Patient Blood Cells Revealed by Flow Cytometry.

<p>PBMC of the younger brother (patient 2) were stained intracellularly with B9 monoclonal antibody to WASP and, where indicated, were stained with surface marker antibodies. (A) Monocytes and lymphocytes. Shown are patient monocytes (left), total lymphocytes (middle), and T lymphocytes (CD3⁺ cells) (right) indicated by filled histograms; cells of a normal individual are indicated by black outlines and isotype control by green outlines. (B) T and NK lymphocytes. CD4 (CD4⁺) and CD8 (CD8^bright) cells, and NK cells (CD56⁺CD3⁻) stained for WASP and, where indicated, for CD45RA to distinguish T cell subsets. Numbers within quadrants indicate the percentage of cells. (C) B-lymphocytes. B cells (CD19⁺) were stained with isotype control antibody (left) or WASP antibody (middle, right) and for CD27 to distinguish B-cell subsets (right). (D) Shown are the percent WASP-positive cells in different populations averaged across the two brothers. ND, not determined.</p

Somatic <i>WASP</i> Alleles.

a<p>Nucleotides and amino acids are numbered relative to normal <i>WASP</i>. Amino acids are identified by single letter code. del, deletion; ins, insertion; fs, frameshift.</p>b<p>Changes of gene and cDNA length (Δ Gene length) are expressed relative to the patients' germ line allele.</p>c<p>Clones indicated by superscript^G were generated from genomic DNA; those indicated by superscript^R were generated from reverse-transcribed RNA. Except where noted (# 3, 6 and 7), each allele was cloned once.</p