A multifunctional network of basic residues confers unique properties to protein kinase CK2

Protein kinase CK2 is characterized by a number of features, including substrate specificity, inhibition by polyanionic compounds and intrasteric down-regulation by its -subunit, which denote a special aptitude to interact with negatively charged ligands. This situation may reflect the presence in CK2 catalytic subunits of several basic residues that are not conserved in the majority of other protein kinases. Some of these residues, notably K49 in the Gly rich loop, K74, K75, K76, K77, K79, R80, K83 in the Lys rich segment and R191, R195, K198 in the p+1 loop, have been shown by mutational studies to be implicated to various extents and with distinct roles in substrate recognition, inhibition by heparin and by pseudosubstrate and instrasteric regulation. Molecular modelization based on crystallographic data provide a rationale for the biochemical observations, showing that several of these basic residues are clustered around the active site where they make contact with individual acidic residues of the peptide substrate. They can also mediate the effect of polyanionic inhibitors (e.g. heparin) and of regulatory elements present in the b-subunit, in the N terminal segment of the catalytic subunit and possibly in other proteins interacting with CK2. Our data also disclose a unique mode of binding of the phosphoacceptor substrate which bridges across the catalytic cleft making contacts with both the lower and upper lobes of CK2.     

A metabolomics-based approach for non-invasive diagnosis of chromosomal anomalies

Introduction

Chromosomal anomalies (CA) are the most frequent fetal anomalies.

Objective

To evaluate the diagnostic performance of a machine learning ensemble model based on the maternal serum metabolomic fingerprint of fetal aneuploidies during the second trimester .

Methods

This is a case-control pilot study. Metabolomic profiles have been obtained on serum of 328 mothers (220 controls and 108 cases), using gas chromatography coupled to mass spectrometry. Eight machines learning and classification models were built and optimized. An ensemble model was built using a voting scheme. All samples were randomly divided into two sets. One was used as training set, the other one for diagnostic performance assessment.

Results

Ensemble machine learning model correctly classified all cases and controls. The accuracy was the same for trisomy 21 and 18; also, the other CA were correctly detected. Elaidic, stearic, linolenic, myristic, benzoic, citric and glyceric acid, mannose, 2-hydroxy butyrate, phenylalanine, proline, alanine and 3-methyl histidine were selected as the most relevant metabolites in class separation.

Conclusion

The proposed model, based on the maternal serum metabolomic fingerprint of fetal aneuploidies during the second trimester, correctly identifies all the cases of chromosomal abnormalities. Overall, this preliminary analysis appeared suggestive of a metabolic environment conductive to increased oxidative stress and a disturbance in the fetal central nervous system development. Maternal serum metabolomics can be a promising tool in the screening of chromosomal defects. Moreover, metabolomics allows to extend our knowledge about biochemical alterations caused by aneuploidies and responsible for the observed phenotypes.

     

Dinoflagellate cyst production at a coastal Mediterranean site

To assess the diversity and seasonality of dinoflagellate cystproduction, surface sediment and trap samples were studied inthe Gulf of Naples (Mediterranean Sea). A total of 59 differentcyst morphotypes were recorded. At the stations within the 70m isobath, sediment assemblages were dominated by calcareousPeridiniales (66–79%), while at the deepest stations non-calcareousPeri-diniales attained the highest percentages (40–49%).The sediment trap sampling, carried out fortnightly over twoannual cycles, revealed high production rates (up to 1.7 x 10⁶cysts m^–2 day^–1) from spring to late autumn of bothyears, with a distinct seasonal production pattern. Althoughrather similar in species composition, the total cyst flux differedmarkedly between the 2 years (1.26 and 0.55 x 108 cysts m^–2year^–1, respectively). Species-specific production patternswere observed: some species formed cysts over several months,others in restricted periods of the year. Cyst-forming speciesconstituted a small part of the planktonic dinoflagellate populationsrecorded in the area. A coupling between the trap material andsurface water plankton was observed for calcareous Peridiniales.This sampling approach allowed the detection of some speciesnever recorded before in the gulf, including two potentiallytoxic species: Alexandrium andersoni and Gymnodinium catenatum-likespecies.     

Nasal immunization of mice with Lactobacillus casei expressing the pneumococcal surface protein C primes the immune system and decreases pneumococcal nasopharyngeal colonization in mice

Streptococcus pneumoniae colonizes the upper respiratory tract of healthy individuals, from where it can be transmitted to the community. Occasionally, bacteria invade sterile niches, causing diseases. The pneumococcal surface protein C (PspC) is a virulence factor that is important during colonization and the systemic phases of the diseases. Here, we have evaluated the effect of nasal or sublingual immunization of mice with Lactobacillus casei expressing PspC, as well as prime-boosting protocols using recombinant PspC, on nasopharyngeal pneumococcal colonization. None of the protocols tested was able to elicit significant levels of anti-PspC antibodies before challenge. However, a significant decrease in pneumococcal recovery from the nasopharynx was observed in animals immunized through the nasal route with L. casei-PspC. Immune responses evaluated after colonization challenge in this group of mice were characterized by an increase in mucosal anti-PspC immunoglobulin A (IgA) 5 days later, a time point in which the pneumococcal loads were already low. A negative correlation between the concentrations of anti-PspC IgA and pneumococcal recovery from the nasopharynx was observed, with animals with the lowest colonization levels having higher IgA concentrations. These results show that nasal immunization with L. casei-PspC primes the immune system of mice, prompting faster immune responses that result in a decrease in pneumococcal colonization.     

In silico comparisons of lipid-related genes between Mycobacterium tuberculosis and BCG vaccine strains

Despite highly variable efficacy, BCG (Bacillus Calmette-Guérin) is the only vaccine available to prevent the tuberculosis (TB). Genomic heterogeneity between attenuated BCG strains and virulent Mycobacterium tuberculosis might help to explain this vaccine’s impaired capacity to induce long-term protection. Here, we investigate the lipid-related genes absent in attenuated BCG strains in order to correlate changes in both lipid metabolism and cell-wall lipid content to vaccine impairment. Whole genome sequences of M. tuberculosis H37Rv and the six most used BCG strains worldwide were aligned and the absent regions functionally categorized. Genomes of the BCG strains showed a total of 14 non-homologous lipid-related genes, including those belonging to mce3 operon, as well as the gene echaA1, which encodes an enoyl-CoA hydratase, and the genes encoding phospholipases PlcA, PlcB and PlcC. Taken together, the depletion of these M. tuberculosis H37Rv genomic regions were associated with marked alterations in lipid-related genes of BCG strains. Such alterations may indicate a dormant-like state and can be determining factors to the vaccine’s inability to induce long-term protection. These lipids can be further evaluated as an adjuvant to boost the current BCG-based vaccine.     

Glycogen synthase kinase-3beta facilitates staurosporine- and heat shock-induced apoptosis. Protection by lithium

The potential role of glycogen synthase kinase-3beta in modulating apoptosis was examined in human SH-SY5Y neuroblastoma cells. Staurosporine treatment caused time- and concentration-dependent increases in the activities of caspase-3 and caspase-9 but not caspase-1, increased proteolysis of poly(ADP-ribose) polymerase, and induced morphological changes consistent with apoptosis. Overexpression of glycogen synthase kinase-3beta to levels 3.5 times that in control cells did not alter basal indices of apoptosis but potentiated staurosporine-induced activation of caspase-3, caspase-9, proteolysis of poly(ADP-ribose) polymerase, and morphological changes indicative of apoptosis. Inhibition of glycogen synthase kinase-3beta by lithium attenuated the enhanced staurosporine-induced activation of caspase-3 in cells overexpressing glycogen synthase kinase-3beta. In cells subjected to heat shock, caspase-3 activity was more than three times greater in glycogen synthase kinase-3beta-transfected than control cells, and this potentiated response was inhibited by lithium treatment. Thus, glycogen synthase kinase-3beta facilitated apoptosis induced by two experimental paradigms. These findings indicate that glycogen synthase kinase-3beta may contribute to pro-apoptotic-signaling activity, that inhibition of glycogen synthase kinase-3beta can contribute to anti-apoptotic-signaling mechanisms, and that the neuroprotective actions of lithium may be due in part to its inhibitory modulation of glycogen synthase kinase-3beta.     

Hydrophobic Derivatives of Glycopeptide Antibiotics as Inhibitors of Protein Kinases

As key regulators of cell signaling, protein kinases (PKs) are attractive targets for therapeutic intervention in a variety of diseases. Herein, we report for the first time the inhibitory activity of polycyclic peptides, particularly, derivatives of glycopeptide antibiotics teicoplanin and eremomycin, against a panel of 12 recombinant human protein kinases and two protein kinases (CK1 and CK2) isolated from rat liver. Several of the investigated compounds inhibited various PKs with IC₅₀ values below 10 μM and caused >90% suppression of the enzyme activity at 10 μM concentration. Kinetic analysis of the protein kinase CK2α inhibition by the teicoplanin aglycon analogue (7) demonstrated the non-competitive mechanism of inhibition (with regard to ATP). Interestingly, the inhibitory activity of some investigated compounds correlated with the earlier described antiviral activity against HIV, HCV, and other corona- and flaviviruses.     

Dietary,Cultural, and Pathogens-Related Selective Pressures Shaped Differential Adaptive Evolution among Native Mexican Populations

Native American genetic ancestry has been remarkably implicated with increased risk of diverse health issues in several Mexican populations, especially in relation to the dramatic changes in environmental, dietary, and cultural settings they have recently undergone. In particular, the effects of these ecological transitions and Westernization of lifestyles have been investigated so far predominantly on Mestizo individuals. Nevertheless, indigenous groups, rather than admixed Mexicans, have plausibly retained the highest proportions of genetic components shaped by natural selection in response to the ancient milieu experienced by Mexican ancestors during their pre-Columbian evolutionary history. These formerly adaptive variants have the potential to represent the genetic determinants of some biological traits that are peculiar to Mexican people, as well as a reservoir of loci with possible biomedical relevance. To test such a hypothesis, we used genome-wide genotype data to infer the unique adaptive evolution of Native Mexican groups selected as reasonable descendants of the main pre-Columbian Mexican civilizations. A combination of haplotype-based and gene-network analyses enabled us to detect genomic signatures ascribable to polygenic adaptive traits plausibly evolved by the main genetic clusters of Mexican indigenous populations to cope with local environmental and/or cultural conditions. Some of these adaptations were found to play a role in modulating the susceptibility/resistance of these groups to certain pathological conditions, thus providing new evidence that diverse selective pressures have contributed to shape the current biological and disease-risk patterns of present-day Native and Mestizo Mexican populations.     

Histamine pharmacokinetics in tumor and host tissues after bolus-dose administration in the rat

In this study, we estimated interstitial histamine concentrations in normal and malignant tissues after a single intravenous (i.v.) injection of 0.5 mg/kg histamine dihydrochloride in the rat. The microdialysis technique was used to collect interstitial fluid from subcutis, liver and a NGW adenocarcinoma. Histamine was absorbed with equal efficiency to all tissues (t 1/2 AB 3.9-7.7 minutes) but maximum concentration (Cmax; nmol/l) of histamine was higher in liver (2,388 +/- 357) than in subcutis (951 +/- 125) (p < 0.01) and subcutaneous tumor (523 +/- 140) (p = 0.01) and, moreover, Cmax in liver tumor (1,752 +/- 326) was higher than in subcutaneous tumor (p = 0.01). The tl/2 elimination was significantly longer in subcutis and subcutaneous tumor than in liver and liver tumor. Area under the curve (AUC; mmol-min/l) for histamine was significantly lower in subcutaneous tumor (9.8 +/- 2.3) than in liver (17.6 +/- 1.9) (p = 0.03) and liver tumor (15.8 +/- 1.8) (p = 0.03). Local tissue blood flow as assessed by the 14C-ethanol method was not significantly altered by the histamine administration. In conclusion, after an i.v. injection of histamine dihydrochloride a higher maximum concentration and AUC of histamine was reached in liver and liver tumor than in subcutaneous tissues.     

Echocardiographic assessment and percutaneous closure of multiple atrial septal defects

Percutaneous coronary intervention can be associated with distal embolization of thrombotic material causing myocardial necrosis and infarction. We discuss the role of intravascular imaging to guide the use of a distal protection device by describing the outcome of a young woman presenting with non-ST elevation myocardial infarction. Coronary angiography demonstrated an isolated minor stenosis in the proximal left anterior descending coronary artery with slight haziness beyond the lesion. Intravascular ultrasound confirmed an extensive thrombus overlying a bulky atherosclerotic plaque. A distal filter wire was therefore successfully used to reduce the risk of distal embolization. The use of intravascular ultrasound in patients presenting with acute coronary syndrome may reveal large thrombi that are difficult to image using conventional angiographic techniques. Intravascular ultrasound can therefore be used as a tool to select lesions requiring distal protection.