The promoter of filamentation (POF1) protein from <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> is an ATPase involved in the protein quality control process

Background

The gene YCL047C, which has been renamed promoter of filamentation gene (POF1), has recently been described as a cell component involved in yeast filamentous growth. The objective of this work is to understand the molecular and biological function of this gene.

Results

Here, we report that the protein encoded by the POF1 gene, Pof1p, is an ATPase that may be part of the Saccharomyces cerevisiae protein quality control pathway. According to the results, Δpof1 cells showed increased sensitivity to hydrogen peroxide, tert-butyl hydroperoxide, heat shock and protein unfolding agents, such as dithiothreitol and tunicamycin. Besides, the overexpression of POF1 suppressed the sensitivity of Δpct1, a strain that lacks a gene that encodes a phosphocholine cytidylyltransferase, to heat shock. In vitro analysis showed, however, that the purified Pof1p enzyme had no cytidylyltransferase activity but does have ATPase activity, with catalytic efficiency comparable to other ATPases involved in endoplasmic reticulum-associated degradation of proteins (ERAD). Supporting these findings, co-immunoprecipitation experiments showed a physical interaction between Pof1p and Ubc7p (an ubiquitin conjugating enzyme) in vivo.

Conclusions

Taken together, the results strongly suggest that the biological function of Pof1p is related to the regulation of protein degradation.

     

Sampling properties of DNA sequence data in phylogenetic analysis

We inferred phylogenetic trees from individual genes and random samples of nucleotides from the mitochondrial genomes of 10 vertebrates and compared the results to those obtained by analyzing the whole genomes. Individual genes are poor samples in that they infrequently lead to the whole-genome tree. A large number of nucleotide sites is needed to exactly determine the whole-genome tree. A relatively small number of sites, however, often results in a tree close to the whole-genome tree. We found that blocks of contiguous sites were less likely to lead to the whole-genome tree than samples composed of sites drawn individually from throughout the genome. Samples of contiguous sites are not representative of the entire genome, a condition that violates a basic assumption of the bootstrap method as it is applied in phylogenetic studies.      

1H NMR studies on bovine cyclophilin: preliminary structural characterization of this specific cyclosporin A binding protein

High-field 1H NMR spectroscopy has been used to study the conformation of the cytosolic cyclosporin A binding protein cyclophilin. For the drug-free form of cyclophilin, spectral editing methods in conjunction with a pH titration were used to identify all four His residues present in the protein, and two-dimensional COSY and RELAY spectroscopy was used to elucidate the scalar connectivities in the aromatic and upfield methyl regions of the spectrum. From these scalar connectivities, it was possible to distinguish between inter- and intraresidue dipolar interactions within the aromatic and upfield methyl regions of cyclophilin in the NOESY spectrum. The results of this analysis showed extensive interresidue cross-relaxation among and between these latter spectral regions indicative of the proximal relationships of several of these residues and the presence of a hydrophobic core within cyclophilin.     

Genotypes and their interaction effects on reproduction and mating‐induced immune activation in Drosophila melanogaster

Mating causes considerable alterations in female physiology and behaviour, and immune gene expression, partly due to proteins transferred from males to females during copulation. The magnitude of these phenotypic changes could be driven by the genotypes of males and females, as well as their interaction. To test this, we carried out a series of genotype‐by‐genotype (G × G) experiments using Drosophila melanogaster populations from two distant geographical locations. We expected lines to have diverged in male reproductive traits and females to differ in their responses to these traits. We examined female physiological and behavioural post‐mating responses to male mating traits, that is behaviour and ejaculate composition, in the short to mid‐term (48 hr) following mating. We then explored whether a sexually transferred molecule, sex peptide (SP), is the mechanism behind our observed female post‐mating responses. Our results show that the genotypes of both sexes as well as the interaction between male and female genotypes affect mating and post‐mating reproductive traits. Immune gene expression of three candidate genes increased in response to mating and was genotype‐dependent but did not show a G × G signature. Males showed genotype‐dependent SP expression in the 7 days following eclosion, but female genotypes showed no differential sensitivity to the receipt of SP. The two genotypes demonstrated clear divergence in physiological traits in short‐ to mid‐term responses to mating, but the longer‐term consequences of these initial dynamics remain to be uncovered.     

Novel Methods for Analysing Bacterial Tracks Reveal Persistence in Rhodobacter sphaeroides

Tracking bacteria using video microscopy is a powerful experimental approach to probe their motile behaviour. The trajectories obtained contain much information relating to the complex patterns of bacterial motility. However, methods for the quantitative analysis of such data are limited. Most swimming bacteria move in approximately straight lines, interspersed with random reorientation phases. It is therefore necessary to segment observed tracks into swimming and reorientation phases to extract useful statistics. We present novel robust analysis tools to discern these two phases in tracks. Our methods comprise a simple and effective protocol for removing spurious tracks from tracking datasets, followed by analysis based on a two-state hidden Markov model, taking advantage of the availability of mutant strains that exhibit swimming-only or reorientating-only motion to generate an empirical prior distribution. Using simulated tracks with varying levels of added noise, we validate our methods and compare them with an existing heuristic method. To our knowledge this is the first example of a systematic assessment of analysis methods in this field. The new methods are substantially more robust to noise and introduce less systematic bias than the heuristic method. We apply our methods to tracks obtained from the bacterial species Rhodobacter sphaeroides and Escherichia coli. Our results demonstrate that R. sphaeroides exhibits persistence over the course of a tumbling event, which is a novel result with important implications in the study of this and similar species.     

Economic evaluation of water loss saving due to the biological control of water hyacinth at New Year’s Dam,Eastern Cape province,South Africa

Water hyacinth Eichhornia crassipes is considered the most damaging aquatic weed in the world. However, few studies have quantified the impact of this weed economically and ecologically, and even fewer studies have quantified the benefits of its control. This paper focuses on water loss saving as the benefit derived from biological control of this plant between 1990 and 2013 at New Year’s Dam, Alicedale, Eastern Cape, South Africa. Estimates of water loss due to evapotranspiration from water hyacinth vary significantly; therefore, the study used three different rates, high, medium and low. A conservative raw agriculture value of R 0.26 per m³ was used to calculate the benefits derived by the water saved. The present benefit and cost values were determined using 10% and 5% discount rates. The benefit/cost ratio at the low evapotranspiration rate was less than one, implying that biological control was not economically viable but, at the higher evapotranspiration rates, the return justified the costs of biological control. However, at the marginal value product of water, the inclusion of the costs of damage to infrastructure, or the adverse effects of water hyacinth on biodiversity, would justify the use of biological control, even at the low transpiration rate.     

The effect of diet and time after bacterial infection on fecundity,resistance, and tolerance in Drosophila melanogaster

Mounting and maintaining an effective immune response in the face of infection can be costly. The outcome of infection depends on two host immune strategies: resistance and tolerance. Resistance limits pathogen load, while tolerance reduces the fitness impact of an infection. While resistance strategies are well studied, tolerance has received less attention, but is now considered to play a vital role in host–pathogen interactions in animals. A major challenge in ecoimmunology is to understand how some hosts maintain their fitness when infected while others succumb to infection, as well as how extrinsic, environmental factors, such as diet, affect defense. We tested whether dietary restriction through yeast (protein) limitation affects resistance, tolerance, and fecundity in Drosophila melanogaster. We predicted that protein restriction would reveal costs of infection. Because infectious diseases are not always lethal, we tested resistance and tolerance using two bacteria with low lethality: Escherichia coli and Lactococcus lactis. We then assayed fecundity and characterized bacterial infection pathology in individual flies at two acute phase time points after infection. As expected, our four fecundity measures all showed a negative effect of a low‐protein diet, but contrary to predictions, diet did not affect resistance to either bacteria species. We found evidence for diet‐induced and time‐dependent variation in host tolerance to E. coli, but not to L. lactis. Furthermore, the two bacteria species exhibited remarkably different infection profiles, and persisted within the flies for at least 7 days postinfection. Our results show that acute phase infections do not necessarily lead to fecundity costs despite high bacterial loads. The influence of intrinsic variables such as genotype are the prevailing factors that have been studied in relation to variation in host tolerance, but here we show that extrinsic factors should also be considered for their role in influencing tolerance strategies.     

Monitoring cancer prognosis,diagnosis and treatment efficacy using metabolomics and lipidomics

Introduction

Cellular metabolism is altered during cancer initiation and progression, which allows cancer cells to increase anabolic synthesis, avoid apoptosis and adapt to low nutrient and oxygen availability. The metabolic nature of cancer enables patient cancer status to be monitored by metabolomics and lipidomics. Additionally, monitoring metabolic status of patients or biological models can be used to greater understand the action of anticancer therapeutics.

Objectives

Discuss how metabolomics and lipidomics can be used to (i) identify metabolic biomarkers of cancer and (ii) understand the mechanism-of-action of anticancer therapies. Discuss considerations that can maximize the clinical value of metabolic cancer biomarkers including case–control, prognostic and longitudinal study designs.

Methods

A literature search of the current relevant primary research was performed.

Results

Metabolomics and lipidomics can identify metabolic signatures that associate with cancer diagnosis, prognosis and disease progression. Discriminatory metabolites were most commonly linked to lipid or energy metabolism. Case–control studies outnumbered prognostic and longitudinal approaches. Prognostic studies were able to correlate metabolic features with future cancer risk, whereas longitudinal studies were most effective for studying cancer progression. Metabolomics and lipidomics can help to understand the mechanism-of-action of anticancer therapeutics and mechanisms of drug resistance.

Conclusion

Metabolomics and lipidomics can be used to identify biomarkers associated with cancer and to better understand anticancer therapies.