Evolution of increased complexity in a molecular machine

Many cellular processes are carried out by molecular 'machines'-assemblies of multiple differentiated proteins that physically interact to execute biological functions. Despite much speculation, strong evidence of the mechanisms by which these assemblies evolved is lacking. Here we use ancestral gene resurrection and manipulative genetic experiments to determine how the complexity of an essential molecular machine--the hexameric transmembrane ring of the eukaryotic V-ATPase proton pump--increased hundreds of millions of years ago. We show that the ring of Fungi, which is composed of three paralogous proteins, evolved from a more ancient two-paralogue complex because of a gene duplication that was followed by loss in each daughter copy of specific interfaces by which it interacts with other ring proteins. These losses were complementary, so both copies became obligate components with restricted spatial roles in the complex. Reintroducing a single historical mutation from each paralogue lineage into the resurrected ancestral proteins is sufficient to recapitulate their asymmetric degeneration and trigger the requirement for the more elaborate three-component ring. Our experiments show that increased complexity in an essential molecular machine evolved because of simple, high-probability evolutionary processes, without the apparent evolution of novel functions. They point to a plausible mechanism for the evolution of complexity in other multi-paralogue protein complexes.     

Intracellular bacterial growth is controlled by a kinase network around PKB/AKT1

With the emergence of multidrug resistant (MDR) bacteria, it is imperative to develop new intervention strategies. Current antibiotics typically target pathogen rather than host-specific biochemical pathways. Here we have developed kinase inhibitors that prevent intracellular growth of unrelated pathogens such as Salmonella typhimurium and Mycobacterium tuberculosis. An RNA interference screen of the human kinome using automated microscopy revealed several host kinases capable of inhibiting intracellular growth of S. typhimurium. The kinases identified clustered in one network around AKT1 (also known as PKB). Inhibitors of AKT1 prevent intracellular growth of various bacteria including MDR-M. tuberculosis. AKT1 is activated by the S. typhimurium effector SopB, which promotes intracellular survival by controlling actin dynamics through PAK4, and phagosome-lysosome fusion through the AS160 (also known as TBC1D4)-RAB14 pathway. AKT1 inhibitors counteract the bacterial manipulation of host signalling processes, thus controlling intracellular growth of bacteria. By using a reciprocal chemical genetics approach, we identified kinase inhibitors with antibiotic properties and their host targets, and we determined host signalling networks that are activated by intracellular bacteria for survival.     

Distribution of phenylalanine transaminase and phenylalanine ammonia-lyase activities in etiolated and light irradiated radish seedlings (Raphanus sativus L.).

In dark-grown Raphanus sativus seedlings the level of phenylalanine transaminase is higher in cotyledons than in root and hypocotyl. The maximum activity of phenylalanine ammonia-lyase (PAL) is found in the root. Only PAL is significantly increased by light.     

Disruptive coloration and background pattern matching

Effective camouflage renders a target indistinguishable from irrelevant background objects. Two interrelated but logically distinct mechanisms for this are background pattern matching (crypsis) and disruptive coloration: in the former, the animal's colours are a random sample of the background; in the latter, bold contrasting colours on the animal's periphery break up its outline. The latter has long been proposed as an explanation for some apparently conspicuous coloration in animals, and is standard textbook material. Surprisingly, only one quantitative test of the theory exists, and one experimental test of its effectiveness against non-human predators. Here we test two key predictions: that patterns on the body's outline should be particularly effective in promoting concealment and that highly contrasting colours should enhance this disruptive effect. Artificial moth-like targets were exposed to bird predation in the field, with the experimental colour patterns on the 'wings' and a dead mealworm as the edible 'body'. Survival analysis supported the predictions, indicating that disruptive coloration is an effective means of camouflage, above and beyond background pattern matching.     

A structural basis for allosteric control of DNA recombination by lambda integrase

Site-specific DNA recombination is important for basic cellular functions including viral integration, control of gene expression, production of genetic diversity and segregation of newly replicated chromosomes, and is used by bacteriophage lambda to integrate or excise its genome into and out of the host chromosome. lambda recombination is carried out by the bacteriophage-encoded integrase protein (lambda-int) together with accessory DNA sites and associated bending proteins that allow regulation in response to cell physiology. Here we report the crystal structures of lambda-int in higher-order complexes with substrates and regulatory DNAs representing different intermediates along the reaction pathway. The structures show how the simultaneous binding of two separate domains of lambda-int to DNA facilitates synapsis and can specify the order of DNA strand cleavage and exchange. An intertwined layer of amino-terminal domains bound to accessory (arm) DNAs shapes the recombination complex in a way that suggests how arm binding shifts the reaction equilibrium in favour of recombinant products.     

Quality in Action Research: Reflections for Second-Order Inquiry

One of the current debates in action research concerns the quality of these practices. Up to now, many contributions have focused on defining specific criteria based on action research epistemology. This article sustains (1) that prior to dealing with these questions, it is necessary to define for what purpose and for whom we are making the evaluation; (2) that this leads us to make a distinction between different evaluation models; and (3) that the quality strategies and criteria will be different for each model. In particular, the article confronts an academic evaluation model as a form of external control over the quality of action research and an internal, participatory evaluation model as a quality strategy aimed at establishing feedback for the process. Final considerations are given about the implications of both models for academia.     

Relative and seasonal abundance of three bark beetle predators (Coleoptera: Trogositidae, Cleridae) across an elevation gradient in ponderosa pine forests of north central Arizona

We examined abundance and flight periodicity of 3 predators of bark beetles (Coleoptera: Curculionidae, Scolytinae), Temnochila chlorodia (Mannerheim) (Coleoptera: Trogositidae), Enoclerus sphegeus (Fabricius) (Coleoptera: Cleridae), and E. lecontei (Wolcott) (Coleoptera: Cleridae), across an elevational gradient of ponderosa pine ( Pinus ponderosa Lawson) forests in north central Arizona. Predator populations were estimated at 10 sites in each of 3 elevation bands (low: 1600–1736 m; mid: 2058–2230 m; high: 2505–2651 m) for 3 years (2004–2006) using pheromone-baited funnel traps targeting 3 primary bark beetle species. We also investigated how predator abundance and flight seasonality related to those of 5 bark beetle species: Ips pini (Say), I. lecontei Swaine, Dendroctonus frontalis Zimmermann, D. brevicomis LeConte, and D. adjunctus Blandford. Temnochila chlorodia was most abundant in the low- and mid-elevation bands, whereas E. sphegeus was most abundant in the high-elevation band. Enoclerus lecontei showed no consistent elevational trend in abundance. Within each elevation band, changes in annual abundance of pooled predator species tracked shifts in abundance of pooled bark beetle species. In general, predator flight initiation coincided with or closely followed bark beetle flight initiation in the spring, but predator flight terminated before flight activity ended for most bark beetle species in the fall. In addition, the ratio of prey to predators was lowest in the summer and highest in the fall. This suggests that all bark beetle species examined may be provided temporal escape from their predators in the fall. For all 3 predator species, the pheromone-baited trap targeting D. brevicomis was less attractive than the pheromone-baited traps targeting I. pini and I. lecontei.     

Additive Biclustering: A Comparison of One New and Two Existing ALS Algorithms

The additive biclustering model for two-way two-mode object by variable data implies overlapping clusterings of both the objects and the variables together with a weight for each bicluster (i.e., a pair of an object and a variable cluster). In the data analysis, an additive biclustering model is fitted to given data by means of minimizing a least squares loss function. To this end, two alternating least squares algorithms (ALS) may be used: (1) PENCLUS, and (2) Baier’s ALS approach. However, both algorithms suffer from some inherent limitations, which may hamper their performance. As a way out, based on theoretical results regarding optimally designing ALS algorithms, in this paper a new ALS algorithm will be presented. In a simulation study this algorithm will be shown to outperform the existing ALS approaches.     

Prediction of central nervous system embryonal tumour outcome based on gene expression.

Embryonal tumours of the central nervous system (CNS) represent a heterogeneous group of tumours about which little is known biologically, and whose diagnosis, on the basis of morphologic appearance alone, is controversial. Medulloblastomas, for example, are the most common malignant brain tumour of childhood, but their pathogenesis is unknown, their relationship to other embryonal CNS tumours is debated, and patients' response to therapy is difficult to predict. We approached these problems by developing a classification system based on DNA microarray gene expression data derived from 99 patient samples. Here we demonstrate that medulloblastomas are molecularly distinct from other brain tumours including primitive neuroectodermal tumours (PNETs), atypical teratoid/rhabdoid tumours (AT/RTs) and malignant gliomas. Previously unrecognized evidence supporting the derivation of medulloblastomas from cerebellar granule cells through activation of the Sonic Hedgehog (SHH) pathway was also revealed. We show further that the clinical outcome of children with medulloblastomas is highly predictable on the basis of the gene expression profiles of their tumours at diagnosis.