Photo-induced protein-RNA cross-linking in mammalian 60-S ribosomal subunits.

Rat liver 60-S ribosomal subunits were submitted to increasing doses of radiation (253.7 nm), at 4 degrees C and 25 degrees C, as previously reported fro 40-S subunits. The existence of protein-RNA cross-linking was demonstrated by two methods. The first consisted in the separation of protein-RNA complex; the second was indirect, and took into account alteration either in the electrophoretic mobility of cross-linked proteins or the separability of 28-S RNA in a 4 M urea/3 M LiCl buffer. The peptide synthetase activity and the sedimentation characteristics of the particles irradiated at 4 degrees C were well preserved, but at 25 degrees C the large subunits were progressively inactivated and unfolded for doses higher than 2 x 10(18) quanta. The dose-dependent variations of protein cross-linkage determined by two-dimensional gel electrophoresis allowed us to distinguish those proteins which reacted at the lowest doses with a first-order reaction from those which cross-linked to RNA after a subtle modification of the subunit structure. At 25 degrees C, all proteins became low-dose reactive. The curve obtained for 28-S RNA cross-linkage was similar to that of the total protein moiety, while those obtained fro the 5-S and 5.8-S RNA (which were parallel) suggest a lower reactivity of these RNAs. As a general rule, proteins from the large subunits were more reactive to RNA than those from the small subunits. This could indicate differences in the organisation of the two subunits.     

An increased requirement for methionine by transformed rat liver epithelial cells in vitro

The growth of two normal and four transformed rat liver epithelial cell lines in a methionine-containing medium and a methionine-deficient medium supplemented with homocysteine was examined. The growth rates of the normal cells on the homocysteine-supplemented medium were approximately one-half the growth rates shown by the same cells in the methionine-containing medium. In contrast, three of the four transformed cell lines studied showed virtually no growth on the homocysteine-supplemented medium, although they grew quite rapidly on the methionine-containing medium. The fourth, transformed by N-methyl-N-nitrosourea, was able to grow on the homocysteine-supplemented medium at about one-third the rate as on the methionine-containing medium. Thus, transformed rat liver epithelial cells resemble other malignant cells in their reduced capacity to grow on homocysteine in the absence of methionine.     

The major growth current through lily pollen tubes enters as K+ and leaves as H+

Growing lily (Lilium longiflorum Thunb.) pollen always drive a current into their tubes and out of their grains. The only external ions needed for growth (and the growth current) are K⁺, H⁺, and Ca²⁺. Increases in K⁺ immediately stimulate the current; while decreases in K⁺ immediately inhibit it. Comparable changes in H⁺ have the opposite effect; while those in Ca²⁺ have very little effect. We infer that most of the steady growth current is carried in by a potassium leak and out by a proton pump; but other considerations indicate that a minor, but controlling, component of the inward current consists of calcium ions.     

Signal sequence insufficiency contributes to neurodegeneration caused by transmembrane prion protein

Protein translocation into the endoplasmic reticulum is mediated by signal sequences that vary widely in primary structure. In vitro studies suggest that such signal sequence variations may correspond to subtly different functional properties. Whether comparable functional differences exist in vivo and are of sufficient magnitude to impact organism physiology is unknown. Here, we investigate this issue by analyzing in transgenic mice the impact of signal sequence efficiency for mammalian prion protein (PrP). We find that replacement of the average efficiency signal sequence of PrP with more efficient signals rescues mice from neurodegeneration caused by otherwise pathogenic PrP mutants in a downstream hydrophobic domain (HD). This effect is explained by the demonstration that efficient signal sequence function precludes generation of a cytosolically exposed, disease-causing transmembrane form of PrP mediated by the HD mutants. Thus, signal sequences are functionally nonequivalent in vivo, with intrinsic inefficiency of the native PrP signal being required for pathogenesis of a subset of disease-causing PrP mutations.     

Listeria monocytogenes induces host DNA damage and delays the host cell cycle to promote infection

Listeria monocytogenes (Lm) is a human intracellular pathogen widely used to uncover the mechanisms evolved by pathogens to establish infection. However, its capacity to perturb the host cell cycle was never reported. We show that Lm infection affects the host cell cycle progression, increasing its overall duration but allowing consecutive rounds of division. A complete Lm infectious cycle induces a S-phase delay accompanied by a slower rate of DNA synthesis and increased levels of host DNA strand breaks. Additionally, DNA damage/replication checkpoint responses are triggered in an Lm dose-dependent manner through the phosphorylation of DNA-PK, H2A.X, and CDC25A and independently from ATM/ATR. While host DNA damage induced exogenously favors Lm dissemination, the override of checkpoint pathways limits infection. We propose that host DNA replication disturbed by Lm infection culminates in DNA strand breaks, triggering DNA damage/replication responses, and ensuring a cell cycle delay that favors Lm propagation.     

Implications of Cardiovascular Disease Risk Assessment Using the WHO/ISH Risk Prediction Charts in Rural India

Cardiovascular disease (CVD) risk in India is currently assessed using the World Health Organization/International Society for Hypertension (WHO/ISH) risk prediction charts since no population-specific models exist. The WHO/ISH risk prediction charts have two versions—one with total cholesterol as a predictor (the high information (HI) model) and the other without (the low information (LI) model). However, information on the WHO/ISH risk prediction charts including guidance on which version to use and when, as well as relative performance of the LI and HI models, is limited. This article aims to, firstly, quantify the relative performance of the LI and HI WHO/ISH risk prediction (for WHO-South East Asian Region D) using data from rural India. Secondly, we propose a pre-screening (simplified) point-of-care (POC) test to identify patients who are likely to benefit from a total cholesterol (TC) test, and subsequently when the LI model is preferential to HI model. Analysis was performed using cross-sectional data from rural Andhra Pradesh collected in 2005 with recorded blood cholesterol measurements (N = 1066). CVD risk was computed using both LI and HI models, and high risk individuals who needed treatment(T _HR) were subsequently identified based on clinical guidelines. Model development for the POC assessment of a TC test was performed through three machine learning techniques: Support Vector Machine (SVM), Regularised Logistic Regression (RLR), and Random Forests (RF) along with a feature selection process. Disagreement in CVD risk predicted by LI and HI WHO/ISH models was 14.5% (n = 155; p<0.01) overall and comprised 36 clinically relevant T _HR patients (31% of patients identified as T _HR by using either model). Using two patient-specific parameters (age, systolic blood pressure), our POC assessment can pre-determine the benefit of TC testing and choose the appropriate risk model (out-of-sample AUCs:RF-0.85,SVM-0.84,RLR:0.82 and maximum sensitivity-98%). The identification of patients benefitting from a TC test for CVD risk stratification can aid planning for resource-allocation and save costs for large-scale screening programmes.     

Sequencing two Tyr::CreERT2 transgenic mouse lines

The Cre/loxP system is a powerful tool that has allowed the study of the effects of specific genes of interest in various biological settings. The Tyr::CreER^T2 system allows for the targeted expression and activity of the Cre enzyme in the melanocyte lineage following treatment with tamoxifen, thus providing spatial and temporal control of the expression of specific target genes. Two independent transgenic mouse models, each containing a Tyr::CreER^T2 transgene, have been generated and are widely used to study melanocyte transformation. In this study, we performed whole genome sequencing (WGS) on genomic DNA from the two Tyr::CreER^T2 mouse models and identified their sites of integration in the C57BL/6 genome. Based on these results, we designed PCR primers to accurately, and efficiently, genotype transgenic mice. Finally, we discussed some of the advantages of each transgenic mouse model.     

First partial cranium of Togocetus from Kpogamé (Togo) and the protocetid diversity in the Togolese phosphate basin

Earliest cetaceans (whales) originated from the early Eocene of Indo-Pakistan, but the group dispersed through most of the oceans of the planet by the late middle to late Eocene. This late Eocene global distribution indicates that important dispersal events took place during the middle Eocene (Lutetian), a globally undersampled time interval that is well documented in the Togolese phosphate series. We report here the first discovery of a partial cetacean cranium from middle Eocene deposits of Togo (West Africa). A 3D model of the cranium and teeth was reconstructed in order to reveal hidden anatomical features. The dental and cranial characteristics of the Togolese specimen recall those of protocetid taxa described in Africa, Asia, and North America, but also display significant differences. In particular, we show that the new specimen shares a number of morphological features with the Togolese taxon Togocetus. Such a hypothesis is further supported by a cladistic analysis including 45 taxa and 167 morphological characters, which recovers the new specimen close to Togocetus as the first offshoot of protocetids. Phylogenetic analysis including all the protocetids remains of Kpogamé confirms the singular diversity of the Togolese phosphate basin, and enables to examine potential connections with faunas from contemporaneous localities in Africa.