RNA folding with soft constraints: reconciliation of probing data and thermodynamic secondary structure prediction

Thermodynamic folding algorithms and structure probing experiments are commonly used to determine the secondary structure of RNAs. Here we propose a formal framework to reconcile information from both prediction algorithms and probing experiments. The thermodynamic energy parameters are adjusted using 'pseudo-energies' to minimize the discrepancy between prediction and experiment. Our framework differs from related approaches that used pseudo-energies in several key aspects. (i) The energy model is only changed when necessary and no adjustments are made if prediction and experiment are consistent. (ii) Pseudo-energies remain biophysically interpretable and hold positional information where experiment and model disagree. (iii) The whole thermodynamic ensemble of structures is considered thus allowing to reconstruct mixtures of suboptimal structures from seemingly contradicting data. (iv) The noise of the energy model and the experimental data is explicitly modeled leading to an intuitive weighting factor through which the problem can be seen as folding with 'soft' constraints of different strength. We present an efficient algorithm to iteratively calculate pseudo-energies within this framework and demonstrate how this approach can be used in combination with SHAPE chemical probing data to improve secondary structure prediction. We further demonstrate that the pseudo-energies correlate with biophysical effects that are known to affect RNA folding such as chemical nucleotide modifications and protein binding.     

Defective osteoclastogenesis by IKKbeta-null precursors is a result of receptor activator of NF-kappaB ligand (RANKL)-induced JNK-dependent apoptosis and impaired differentiation

It has been reported previously that inhibitory kappaB kinase (IKK) supports osteoclastogenesis through NF-kappaB-mediated prevention of apoptosis. This finding suggests that the ligand for receptor activator of NF-kappaB (RANKL), the master osteoclastogenic cytokine, induces apoptosis of osteoclast precursors (OCPs) in the absence of IKKbeta/NF-kappaB competency. To validate this hypothesis, we sought to determine the pro-apoptotic signaling factors induced by RANKL in IKKbeta-null osteoclast OCPs and to rescue osteoclast differentiation in the absence of IKKbeta through their inhibition. To accomplish this, we generated mice that lack IKKbeta in multiple hematopoietic lineages, including OCPs. We found that these mice possess both in vitro and in vivo defects in osteoclast generation, in concurrence with previous reports, and that this defect is a result of susceptibility to RANKL-mediated apoptosis as a result of gain-of-function of JNK activation. We demonstrate that differentiation of OCPs depends on IKKbeta because reduced IKKbeta mRNA expression correlates with impaired induction of osteoclast differentiation markers in response to RANKL stimulation. We further show that fine-tuned inhibition of JNK activation in these cells inhibits RANKL-induced apoptosis and restores the ability of IKKbeta-null OCPs to become mature osteoclasts. Our data highlight the pro-osteoclastogenic and anti-apoptotic roles of IKKbeta in OCPs and identify a pro-apoptotic mechanism activated within the RANK signalosome.     

Identification of oxidative stress and Toll-like receptor 4 signaling as a key pathway of acute lung injury

Multiple lung pathogens such as chemical agents, H5N1 avian flu, or SARS cause high lethality due to acute respiratory distress syndrome. Here we report that Toll-like receptor 4 (TLR4) mutant mice display natural resistance to acid-induced acute lung injury (ALI). We show that TLR4-TRIF-TRAF6 signaling is a key disease pathway that controls the severity of ALI. The oxidized phospholipid (OxPL) OxPAPC was identified to induce lung injury and cytokine production by lung macrophages via TLR4-TRIF. We observed OxPL production in the lungs of humans and animals infected with SARS, Anthrax, or H5N1. Pulmonary challenge with an inactivated H5N1 avian influenza virus rapidly induces ALI and OxPL formation in mice. Loss of TLR4 or TRIF expression protects mice from H5N1-induced ALI. Moreover, deletion of ncf1, which controls ROS production, improves the severity of H5N1-mediated ALI. Our data identify oxidative stress and innate immunity as key lung injury pathways that control the severity of ALI.     

Exploring sex differences in diets and activity patterns through dental and skeletal studies in populations from ancient Corinth,Greece

Sex and temporal differences are assessed in relation to dietary habits and activity patterns in three ancient populations from Corinth, Greece. The skeletal sample spans time from the Geometric to the Early Byzantine Period (9th c. BCE-5th c. CE). Dental caries and tooth wear have been proven to be reliable dietary indicators. Similarly, spinal osteoarthritis, spinal facet remodeling and Schmorl’s nodes, have been used to infer activity patterns.     

Periodontal disease in the Mycenean (1450–1150 BC) population of Aghia Triada, W. Peloponnese, Greece

This study investigates the incidence of periodontitis in a Mycenaean population unearthed at the cemetery of Aghia Triada (West Peloponnese, Greece) during the 1989–1997 field season. The material consists of 172 dry skulls. Demographic parameters of sex and age were difficult to be assigned due to the bad state of preservation of the skeletal material. The ratio was 50 males, 48 females, 65 unidentified and 9 children, and we estimated an average age of 38 years. In this work we used traditional and modern methods to determine the incidence of periodontitis in the archeological human dental bone. We also recorded other dental diseases, such as ante mortem tooth loss, caries and attrition. The results showed that periodontitis has affected 35% of the jaws. A notable percentage of the individuals — 24% — lost three or more teeth during their lifetime and a total 53% of the population had extracted teeth before death. This paper points out that the ancient jaws present a high proportion of ante-mortem tooth loss, attrition and deep caries, whereas the frequency of periodontitis does not seem to differ from that of other prehistoric samples.     

Instruments to Assess Secondhand Smoke Exposure in Large Cohorts of Never Smokers: The Smoke Scales

The objectives of this study were to: (i) to develop questionnaires that can identify never-smoking children and adults experiencing increased exposure to secondhand smoke (SHS+), (ii) to determine their validity against hair nicotine, and (iii) assess their reliability. A sample of 191 children (85 males; 106 females; 7–18 years) and 95 adult (23 males; 72 females; 18–62 years) never-smokers consented to hair nicotine analysis and answered a large number of questions assessing all sources of SHS. A randomly-selected 30% answered the questions again after 20–30 days. Prevalence of SHS+ in children and adults was 0.52±0.07 and 0.67±0.10, respectively (p<0.05). The Smoke Scale for Children (SS-C) and the Smoke Scale for Adults (SS-A) were developed via factor analysis and included nine questions each. Positivity criteria for SS-C and SS-A via receiver operating characteristics curve analysis were identified at >16.5 and >16, respectively. Significant Kappa agreement (p<0.05) was confirmed when comparing the SS-C and SS-A to hair nicotine concentration. Reliability analyses demonstrated that the SS-C and SS-A scores obtained on two different days are highly correlated (p<0.001) and not significantly different (p>0.05). Area under the curve and McNemar''s Chi-square showed no pair-wise differences in sensitivity and specificity at the cutoff point between the two different days for SS-C and SS-A (p>0.05). We conclude that the SS-C and the SS-A represent valid, reliable, practical, and inexpensive instruments to identify children and adult never-smokers exposed to increased SHS. Future research should aim to further increase the validity of the two questionnaires.     

SPPS of protected peptidyl aminoalkyl amides.

Monophthaloyl diamines derived from naturally occurring amino acids were attached through their free amino functions to resins of the trityl type. The phthaloyl groups were removed by hydrazinolysis, and peptide chains were assembled using Fmoc/tBu-amino acids on the liberated amino functions. The peptidyl aminoalkyl amides obtained were cleaved from the resins by mild acidolysis, with the tBu-side chain protection remaining intact.