Protein kinase D as a potential new target for cancer therapy

Protein kinase D is a novel family of serine/threonine kinases and diacylglycerol receptors that belongs to the calcium/calmodulin-dependent kinase superfamily. Evidence has established that specific PKD isoforms are dysregulated in several cancer types, and PKD involvement has been documented in a variety of cellular processes important to cancer development, including cell growth, apoptosis, motility, and angiogenesis. In light of this, there has been a recent surge in the development of novel chemical inhibitors of PKD. This review focuses on the potential of PKD as a chemotherapeutic target in cancer treatment and highlights important recent advances in the development of PKD inhibitors.     

Could a dysfunction of ferritin be a determinant factor in the aetiology of some neurodegenerative diseases?

Background

The concentration of iron in the brain increases with aging. Furthermore, it has also been observed that patients suffering from neurological diseases (e.g. Parkinson, Alzheimer…) accumulate iron in the brain regions affected by the disease. Nevertheless, it is still not clear whether this accumulation is the initial cause or a secondary consequence of the disease. Free iron excess may be an oxidative stress source causing cell damage if it is not correctly stored in ferritin cores as a ferric iron oxide redox-inert form.

Scope

Both, the composition of ferritin cores and their location at subcellular level have been studied using analytical transmission electron microscopy in brain tissues from progressive supranuclear palsy (PSP) and Alzheimer disease (AD) patients.

Major conclusions

Ferritin has been mainly found in oligodendrocytes and in dystrophic myelinated axons from the neuropili in AD. In relation to the biomineralization of iron inside the ferritin shell, several different crystalline structures have been observed in the study of physiological and pathological ferritin. Two cubic mixed ferric–ferrous iron oxides are the major components of pathological ferritins whereas ferrihydrite, a hexagonal ferric iron oxide, is the major component of physiological ferritin. We hypothesize a dysfunction of ferritin in its ferroxidase activity.

General significance

The different mineralization of iron inside ferritin may be related to oxidative stress in olygodendrocites, which could affect myelination processes with the consequent perturbation of information transference.     

Solution conformation of a tetradecapeptide stabilized by two di‐n‐propyl glycine residues

The solution conformation of a designed tetradecapeptide Boc‐Val‐Ala‐Leu‐Dpg‐Val‐Ala‐Leu‐Val‐Ala‐Leu‐Dpg‐Val‐Ala‐Leu‐OMe (Dpg‐14) containing two di‐n‐propyl glycine (Dpg) residues has been investigated by ¹H NMR and circular dichroism in organic solvents. The peptide aggregates formed at a concentration of 3 mM in the apolar solvent CDCl₃ were broken by the addition of 12% v/v of the more polar solvent DMSO‐d₆. Successive N_iH N_i+1H NOEs observed over the entire length of the sequence in this solvent mixture together with the observation of several characteristic medium‐range NOEs support a major population of continuous helical conformations for Dpg‐14. Majority of the observed coupling constants ( ) also support ? values in the helical conformation. Circular dichroism spectra recorded in methanol and propan‐2‐ol give further support in favor of helical conformation for Dpg‐14 and the stability of the helix at higher temperature. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd.     

Embodied Resource Flows and Product Flows

We develop the absorbing Markov chain (AMC) for describing in detail the network of paths through an industrial system taken by an embodied resource from extraction through intermediate products and, finally, consumer products. We refer to this as a resource‐specific network. This work builds on a recent literature in industrial ecology that uses an AMC to quantify the number of times a resource passes through a recycling sector before ending up in a landfill. Our objective is to incorporate into that analysis an input‐output (IO) table so that the resource paths explicitly take account of the interdependence of sectors through their reliance on intermediate products. This feature makes it possible to track multiple resources simultaneously and consistently and to represent both resources and products in mixed units. Hypothetical scenarios about technological changes and changes in consumer demand are analyzed with an IO model, and model solutions generate the AMC database. A numerical example is provided. We identify the three most critical enhancements to the standard IO model that will be needed for analyzing material cycles: the incorporation of waste‐processing sectors, stock and flow relationships, and international trade. The idea is to implement an AMC after each modeling step for analyses, such as tracking a resource extracted in one region to landfills in other regions and evaluating ways to intensify secondary recovery at key junctures in between.     

Temperature-dependent structural changes in intrinsically disordered proteins: Formation of α-helices or loss of polyproline II?

Structural characterization of intrinsically disordered proteins (IDPs) is mandatory for deciphering their potential unique physical and biological properties. A large number of circular dichroism (CD) studies have demonstrated that a structural change takes place in IDPs with increasing temperature, which most likely reflects formation of transient α-helices or loss of polyproline II (PPII) content. Using three IDPs, ACTR, NHE1, and Spd1, we show that the temperature-induced structural change is common among IDPs and is accompanied by a contraction of the conformational ensemble. This phenomenon was explored at residue resolution by multidimensional NMR spectroscopy. Intrinsic chemical shift referencing allowed us to identify regions of transiently formed helices and their temperature-dependent changes in helicity. All helical regions were found to lose rather than gain helical structures with increasing temperature, and accordingly these were not responsible for the change in the CD spectra. In contrast, the nonhelical regions exhibited a general temperature-dependent structural change that was independent of long-range interactions. The temperature-dependent CD spectroscopic signature of IDPs that has been amply documented can be rationalized to represent redistribution of the statistical coil involving a general loss of PPII conformations.     

Development and validation of a sensitive and selective UHPLC-MS/MS method for simultaneous determination of both free and total eicosapentaeonic acid and docosahexenoic acid in human plasma

A sensitive, selective, and quantitative method for the simultaneous determination of free and total eicosapentaeonic acid (EPA) and docosahexenoic acid (DHA) has been developed and validated in human plasma using fatty acid free human serum albumin as a surrogate matrix. Clean-up for free EPA and DHA employs a liquid-liquid extraction with hexane to remove plasma interferences and provide for cleaner chromatography. The method for total EPA and DHA requires a digestion of the triglycerides followed by liquid-liquid extraction with hexane. Ultra high performance liquid chromatography (UHPLC) technology on a BEH C18 stationary phase column with 1.7 μm particle size was used for chromatographic separation, coupled to tandem mass spectrometry (UHPLC-MS/MS). The method for free EPA and DHA was validated over the concentration range of 0.05-25 μg/mL, while total EPA and DHA concentration range was 0.5-250 μg/mL. The results from assay validation show that the method is rugged, precise, accurate, and well suited to support pharmacokinetic studies. To our knowledge, this work represents the first UHPLC-MS/MS based method that combines both free and total EPA and DHA with a relatively small sample volume (25 μL aliquot) and a run time of 1.5 min, facilitating automation and high throughput analysis.     

2D-HPLC and MALDI-TOF/TOF analysis of barley proteins glycated during brewing

The barley proteins have been the subject of interests of many research groups dealing with barley grains, malt and beer. The proteins which remain intact after harsh malting conditions influence the quality and flavor of beer. The characteristic feature of the proteins present in malt and beer is their extensive modification with carbohydrates, mainly glucose that comes from the starch degradation during technological processes. The degree of the protein glycation has an effect on the quality of malt and beer and on the properties of the beer foam. A combination of two-dimensional high performance liquid chromatography (2D-HPLC) and matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF/TOF MS) was used for the analysis of the protein extracts that were reduced, alkylated, and degraded enzymatically without prior protein separation. This so-called "shot-gun" approach enabled us to determine glycation sites in one third of the proteins identified in the study and to propose potential glycation markers for fast and efficient monitoring during malting.     

MMP-12 catalytic domain recognizes and cleaves at multiple sites in human skin collagen type I and type III

Collagens of either soft connective or mineralized tissues are subject to continuous remodeling and turnover. Undesired cleavage can be the result of an imbalance between proteases and their inhibitors. Owing to their superhelical structure, collagens are resistant to many proteases and matrix metalloproteinases (MMPs) are required to initiate further degradation by other enzymes. Several MMPs are known to degrade collagens, but the action of MMP-12 has not yet been studied in detail. In this work, the potential of MMP-12 in recognizing sites in human skin collagen types I and III has been investigated. The catalytic domain of MMP-12 binds to the triple helix and cleaves the typical sites -Gly₇₇₅-Leu₇₇₆- in α-2 type I collagen and -Gly₇₇₅-Ile₇₇₆- in α-1 type I and type III collagens and at multiple other sites in both collagen types. Moreover, it was observed that the region around these typical sites contains comparatively less prolines, of which some have been proven to be only partially hydroxylated. This is of relevance since partial hydroxylation in the vicinity of a potential scissile bond may have a local effect on the conformational thermodynamics with probable consequences on the collagenolysis process. Taken together, the results of the present work confirm that the catalytic domain of MMP-12 alone binds and degrades collagens I and III.     

Moesin‐dependent cytoskeleton remodelling is associated with an anaplastic phenotype of pancreatic cancer

Cell motility is controlled by the dynamic cytoskeleton and its related proteins, such as members of the ezrin/radixin/moesin (ERM) family, which act as signalling molecules inducing cytoskeleton remodelling. Although ERM proteins have been identified as important factors in various malignancies, functional redundancy between these proteins has hindered the dissection of their individual contribution. The aim of the present study was to analyse the functional role of moesin in pancreatic malignancies. Cancer cells of different malignant lesions of human and transgenic mice pancreata were evaluated by immunohistochemistry. For functional analysis, cell growth, adhesion and invasion assays were carried out after transient and stable knock‐down of moesin expression in pancreatic cancer cells. In vivo tumourigenicity was determined using orthotopic and metastatic mouse tumour models. We now show that moesin knock‐down increases migration, invasion and metastasis and influences extracellular matrix organization of pancreatic cancer. Moesin‐regulated migratory activities of pancreatic cancer cells were in part promoted through cellular translocation of β‐catenin, and re‐distribution and organization of the cytoskeleton. Analysis of human and different transgenic mouse pancreatic cancers demonstrated that moesin is a phenotypic marker for anaplastic carcinoma, suggesting that this ERM protein plays a specific role in pancreatic carcinogenesis.