Quantitative phosphoproteomic analysis of signaling network dynamics

Protein phosphorylation mediated cellular signaling is a highly regulated, dynamic process that controls many aspects of cellular biology. Over the past few years many methods have been developed to quantify temporal dynamics of protein phosphorylation, including mass spectrometry, which can be applied in both an unbiased, discovery mode and in a targeted mode to monitor specific phosphorylation sites. Other methods, such as kinase activity assays and antibody microarrays, have been applied to quantify central nodes in the signaling network, yielding intriguing biological insights. This review provides a concise overview of the latest advances in the quantitative analysis of signaling dynamics including a brief commentary on the future of the field.     

Semiparametric modeling of longitudinal measurements and time-to-event data--a two-stage regression calibration approach

SUMMARY: In this article we investigate regression calibration methods to jointly model longitudinal and survival data using a semiparametric longitudinal model and a proportional hazards model. In the longitudinal model, a biomarker is assumed to follow a semiparametric mixed model where covariate effects are modeled parametrically and subject-specific time profiles are modeled nonparametrially using a population smoothing spline and subject-specific random stochastic processes. The Cox model is assumed for survival data by including both the current measure and the rate of change of the underlying longitudinal trajectories as covariates, as motivated by a prostate cancer study application. We develop a two-stage semiparametric regression calibration (RC) method. Two variations of the RC method are considered, risk set regression calibration and a computationally simpler ordinary regression calibration. Simulation results show that the two-stage RC approach performs well in practice and effectively corrects the bias from the naive method. We apply the proposed methods to the analysis of a dataset for evaluating the effects of the longitudinal biomarker PSA on the recurrence of prostate cancer.     

Magnitude and direction of DNA bending induced by screw-axis orientation: influence of sequence, mismatches and abasic sites

DNA-bending flexibility is central for its many biological functions. A new bending restraining method for use in molecular mechanics calculations and molecular dynamics simulations was developed. It is based on an average screw rotation axis definition for DNA segments and allows inducing continuous and smooth bending deformations of a DNA oligonucleotide. In addition to controlling the magnitude of induced bending it is also possible to control the bending direction so that the calculation of a complete (2-dimensional) directional DNA-bending map is now possible. The method was applied to several DNA oligonucleotides including A(adenine)-tract containing sequences known to form stable bent structures and to DNA containing mismatches or an abasic site. In case of G:A and C:C mismatches a greater variety of conformations bent in various directions compared to regular B-DNA was found. For comparison, a molecular dynamics implementation of the approach was also applied to calculate the free energy change associated with bending of A-tract containing DNA, including deformations significantly beyond the optimal curvature. Good agreement with available experimental data was obtained offering an atomic level explanation for stable bending of A-tract containing DNA molecules. The DNA-bending persistence length estimated from the explicit solvent simulations is also in good agreement with experiment whereas the adiabatic mapping calculations with a GB solvent model predict a bending rigidity roughly two times larger.     

Phylogenetic classification of short environmental DNA fragments

Metagenomics is providing striking insights into the ecology of microbial communities. The recently developed massively parallel 454 pyrosequencing technique gives the opportunity to rapidly obtain metagenomic sequences at a low cost and without cloning bias. However, the phylogenetic analysis of the short reads produced represents a significant computational challenge. The phylogenetic algorithm CARMA for predicting the source organisms of environmental 454 reads is described. The algorithm searches for conserved Pfam domain and protein families in the unassembled reads of a sample. These gene fragments (environmental gene tags, EGTs), are classified into a higher-order taxonomy based on the reconstruction of a phylogenetic tree of each matching Pfam family. The method exhibits high accuracy for a wide range of taxonomic groups, and EGTs as short as 27 amino acids can be phylogenetically classified up to the rank of genus. The algorithm was applied in a comparative study of three aquatic microbial samples obtained by 454 pyrosequencing. Profound differences in the taxonomic composition of these samples could be clearly revealed.     

Observations of serotonin and FMRFamide-like immunoreactivity in palp sensory structures and the anterior nervous system of spionid polychaetes

Evidence suggests that ciliated sensory structures on the feeding palps of spionid polychaetes may function as chemoreceptors to modulate deposit-feeding activity. To investigate the probable sensory nature of these ciliated cells, we used immunohistochemistry, epi-fluorescence, and confocal laser scanning microscopy to label and image sensory cells, nerves, and their organization relative to the anterior central nervous system in several spionid polychaete species. Antibodies directed against acetylated alphatubulin were used to label the nervous system and detail the innervation of palp sensory cells in all species. In addition, the distribution of serotonin (5-HT) and FMRFamide-like immunoreactivity was compared in the spionid polychaetes Dipolydora quadrilobata and Pygospio elegans. The distribution of serotonin immunoreactivity was also examined in the palps of Polydora cornuta and Streblospio benedicti. Serotonin immunoreactivity was concentrated in cells underlying the food groove of the palps, in the palp nerves, and in the cerebral ganglion. FMRFamide-like immunoreactivity was associated with the cerebral ganglia, nuchal organs and palp nerves, and also with the perikarya of ciliated sensory cells on the palps.     

Biphenylsulfonyl-thiophene-carboxamidine inhibitors of the complement component C1s

Complement activation has been implicated in disease states such as hereditary angioedema, ischemia-reperfusion injury, acute respiratory distress syndrome, and acute transplant rejection. Even though the complement cascade provides several protein targets for potential therapeutic intervention only two complement inhibitors have been approved so far for clinical use including anti-C5 antibodies for the treatment of paroxysmal nocturnal hemoglobinuria and purified C1-esterase inhibitor replacement therapy for the control of hereditary angioedema flares. In the present study, optimization of potency and physicochemical properties of a series of thiophene amidine-based C1s inhibitors with potential utility as intravenous agents for the inhibition of the classical pathway of complement is described.     

Surgical stabilisation of the spine compared with a programme of intensive rehabilitation for the management of patients with chronic low back pain: cost utility analysis based on a randomised controlled trial

Objective To determine whether, from a health provider and patient perspective, surgical stabilisation of the spine is cost effective when compared with an intensive programme of rehabilitation in patients with chronic low back pain.Design Economic evaluation alongside a pragmatic randomised controlled trial.Setting Secondary care.Participants 349 patients randomised to surgery (n = 176) or to an intensive rehabilitation programme (n = 173) from 15 centres across the United Kingdom between June 1996 and February 2002.Main outcome measures Costs related to back pain and incurred by the NHS and patients up to 24 months after randomisation. Return to paid employment and total hours worked. Patient utility as estimated by using the EuroQol EQ-5D questionnaire at several time points and used to calculate quality adjusted life years (QALYs). Cost effectiveness was expressed as an incremental cost per QALY.Results At two years, 38 patients randomised to rehabilitation had received rehabilitation and surgery whereas just seven surgery patients had received both treatments. The mean total cost per patient was estimated to be £7830 (SD £5202) in the surgery group and £4526 (SD £4155) in the intensive rehabilitation arm, a significant difference of £3304 (95% confidence interval £2317 to £4291). Mean QALYs over the trial period were 1.004 (SD 0.405) in the surgery group and 0.936 (SD 0.431) in the intensive rehabilitation group, giving a non-significant difference of 0.068 (–0.020 to 0.156). The incremental cost effectiveness ratio was estimated to be £48 588 per QALY gained (–£279 883 to £372 406).Conclusion Two year follow-up data show that surgical stabilisation of the spine may not be a cost effective use of scarce healthcare resources. However, sensitivity analyses show that this could change—for example, if the proportion of rehabilitation patients requiring subsequent surgery continues to increase.     

UV effects that come and go: a global comparison of marine benthic community level impacts

Ambient UV radiation has substantially increased during the last decades, but its impact on marine benthic communities is hardly known. The aim of this study was to globally compare and quantify how shallow hard‐bottom communities are affected by UV during early succession. Identical field experiments in 10 different coastal regions of both hemispheres produced a consistent but unexpected pattern: (i) UV radiation affected species diversity and community biomass in a very similar manner, (ii) diversity and biomass were reduced to a larger extent by UVA than UVB radiation, (iii) ambient UV levels did not affect the composition of the communities, and (iv) any UV effects disappeared during species succession after 2–3 months. Thus, current levels of UV radiation seem to have small, predictable, and transient effects on shallow marine hard‐bottom communities.     

Catechin and hydroxybenzhydrols as models for the environmental photochemistry of tannins and lignins.

The photochemistry of several model plant-derived compounds has been studied in aqueous solution. In particular, the reactions of catechin as a model tannin and methoxy-substituted hydroxybenzhydrols as model lignin functionalities were investigated. Tannins and lignins constitute a significant portion of the humic substances in aquatic systems, which are themselves the main component of dissolved organic matter thought to be responsible for the absorption and attenuation of light in these environments. Catechin (1) was found to undergo a reversible photoisomerization reaction to give epicatechin (2). Such a reaction is an explicit example of a photon absorbing process that enables catechin (1) and its derivatives to act as natural sunscreens by attenuating light energy through non-destructive reactions. The methoxy-substituted hydroxybenzhydrols were found to undergo photosolvolysis reactions via efficient generation of quinone methide intermediates. The intermediate quinone methides were observed to be longer lived, and thus more stable, than previously studied hydroxybenzhydrol derivatives. The meta-hydroxybenzhydrol isomer (5) was found to undergo additional chemistry leading to the production of a ring-closed fluorene from the quinone methide intermediate.