Bidirectional signaling of mammary epithelium and stroma: implications for breast cancer--preventive actions of dietary factors

The mammary gland is composed of two major cellular compartments: a highly dynamic epithelium that undergoes cycles of proliferation, differentiation and apoptosis in response to local and endocrine signals and the underlying stroma comprised of fibroblasts, endothelial cells and adipocytes, which collectively form the mammary fat pad. Breast cancer originates from subversions of normal growth regulatory pathways in mammary epithelial cells due to genetic mutations and epigenetic modifications in tumor suppressors, oncogenes and DNA repair genes. Diet is considered a highly modifiable determinant of breast cancer risk; thus, considerable efforts are focused on understanding how certain dietary factors may promote resistance of mammary epithelial cells to growth dysregulation. The recent indications that stromal cells contribute to the maintenance of the mammary epithelial ‘niche’ and the increasing appreciation for adipose tissue as an endocrine organ with a complex secretome have led to the novel paradigm that the mammary stromal compartment is itself a relevant target of bioactive dietary factors. In this review, we address the potential influence of dietary factors on mammary epithelial-stromal bidirectional signaling to provide mechanistic insights into how dietary factors may promote early mammary epithelial differentiation to decrease adult breast cancer risk.     

Transformation of Gaeumannomyces graminis with β‐Glucuronidase Reporter and Hygromycin Resistance Genes

Take‐all disease is caused by Gaeumannomyces graminis, (Sacc.) Arx & D. Olivier, a soil‐borne fungus, which colonizes the root and crown tissue of many members of the Poaceae plant family. This fungus is able to grow along the surface of roots as darkly pigmented runner hyphae, which has the ability to penetrate the root. Here, we describe a genetic transformation of G. graminis var. graminis by using polyethylene glycol (PEG)‐based protoplast transformation. Fungus cells were transformed with a plasmid, pHPG, containing the gusA reporter gene that codes for β‐glucuronidase (GUS) and the hph gene for hygromycin resistance as the selectable marker. A de novo transformant selection assay was developed to identify the putative transformants that were expressing the hph gene. In addition, the transformed cells maintained the ability to infect the plant tissues. The GUS‐expressing fungus can be used to study fungal infection processes including fungal penetration, colonization and the role(s) of melanin during pathogenesis. Thus, this study is the first report of G. graminis var. graminis transformed with a visibly detectable reporter gene that provides a useful tool to a better understanding of host–Gaeumannomyces interactions.     

Enantiomers of naringenin as pleiotropic,stereoselective inhibitors of cytochrome P450 isoforms

Interactions between naringenin and the cytochrome P450 (CYP) system have been of interest since the first demonstration that grapefruit juice reduced CYP3A activity. The effects of naringenin on other CYP isoforms have been less investigated. In addition, it is well known that interactions with enzymes are often stereospecific, but due to the lack of readily available pure naringenin enantiomers, the enantioselectivity of its effects has not been characterized. We isolated pure naringenin enantiomers by chiral high‐performance liquid chromatography and tested the ability of (R)‐,(S)‐ and rac‐naringenin to inhibit several important drug‐metabolizing CYP isoforms using recombinant enzymes and pooled human liver microsomes. Naringenin was able to inhibit CYP19, CYP2C9, and CYP2C19 with IC₅₀ values below 5 μM. No appreciable inhibition of CYP2B6 or CYP2D6 was observed at concentrations up to 10 μM. Whereas (S)‐naringenin was 2‐fold more potent as an inhibitor of CYP19 and CYP2C19 than (R)‐naringenin, (R)‐naringenin was 2‐fold more potent for CYP2C9 and CYP3A. Chiral flavanones like naringenin are difficult to separate into their enantiomeric forms, but enantioselective effects may be observed that ultimately impact clinical effects. Inhibition of specific drug metabolizing enzymes by naringenin observed in vitro may be exploited to understand pharmacokinetic changes seen in vivo. Chirality, 2011. © 2011 Wiley‐Liss, Inc.     

Improvement of the production of a red pigment in Penicillium sp. HSD07B synthesized during co-culture with Candida tropicalis

Co-culture of Penicillium sp. HSD07B and Candida tropicalis resulted in the production of a red pigment consisting of six components as determined by TLC and HPLC. The pigment showed no acute toxicity in mice and was mot mutagenic in the Ames test. The pigment was stable between pH 2 and 10 and temperatures of 10-100 °C and exhibited good photo-stability and resistance to oxidization by hydrogen peroxide and reduction by Na₂SO₃. Glucose and ratio of C. tropicalis to strain HSD07B (w/w) in the inoculum were the important factors influencing production of the pigment. Under optimized conditions, a pigment yield of 2.75 and 7.7 g/l was obtained in a shake-flask and a 15 l bioreactor, respectively. Thus, co-culture of strain HSD07B and C. tropicalis is a promising way to produce a red pigment potentially useful for coloring applications.     

An accurate feature-based method for identifying DNA-binding residues on protein surfaces

Proteins that interact with DNA play vital roles in all mechanisms of gene expression and regulation. In order to understand these activities, it is crucial to analyze and identify DNA-binding residues on DNA-binding protein surfaces. Here, we proposed two novel features B-factor and packing density in combination with several conventional features to characterize the DNA-binding residues in a well-constructed representative dataset of 119 protein-DNA complexes from the Protein Data Bank (PDB). Based on the selected features, a prediction model for DNA-binding residues was constructed using support vector machine (SVM). The predictor was evaluated using a 5-fold cross validation on above dataset of 123 DNA-binding proteins. Moreover, two independent datasets of 83 DNA-bound protein structures and their corresponding DNA-free forms were compiled. The B-factor and packing density features were statistically analyzed on these 83 pairs of holo-apo proteins structures. Finally, we developed the SVM model to accurately predict DNA-binding residues on protein surface, given the DNA-free structure of a protein. Results showed here indicate that our method represents a significant improvement of previously existing approaches such as DISPLAR. The observation suggests that our method will be useful in studying protein-DNA interactions to guide consequent works such as site-directed mutagenesis and protein-DNA docking.     

Predicting kinetic constants of protein-protein interactions based on structural properties

Elucidating kinetic processes of protein–protein interactions (PPI) helps to understand how basic building blocks affect overall behavior of living systems. In this study, we used structure‐based properties to build predictive models for kinetic constants of PPI. A highly diverse PPI dataset, protein–protein kinetic interaction data and structures (PPKIDS), was built. PPKIDS contains 62 PPI with complex structures and kinetic constants measured experimentally. The influence of structural properties on kinetics of PPI was studied using 35 structure‐based features, describing different aspects of complex structures. Linear models for the prediction of kinetic constants were built by fitting with selected subsets of structure‐based features. The models gave correlation coefficients of 0.801, 0.732, and 0.770 for k_off, k_on, and K_d, respectively, in leave‐one‐out cross validations. The predictive models reported here use only protein complex structures as input and can be generally applied in PPI studies as well as systems biology modeling. Our study confirmed that different properties play different roles in the kinetic process of PPI. For example, k_on was affected by overall structural features of complexes, such as the composition of secondary structures, the change of translational and rotational entropy, and the electrostatic interaction; while k_off was determined by interfacial properties, such as number of contacted atom pairs per 100 Ų. This information provides useful hints for PPI design. Proteins 2010;79:720–734. © 2010 Wiley‐Liss, Inc.     

Enhancing systematic protein-protein docking methods using ray casting: application to ATTRACT

Systematic protein-protein docking methods need to evaluate a huge number of different probe configurations, thus leading to high computational cost. We present an efficient filter-ray casting filter (RCF)-that enables a notable speed-up of systematic protein-protein docking. The high efficiency of RCF is the outcome of the following factors: (i) extracting of pockets and protrusions on the surfaces of the proteins using visibilities; (ii) a ray casting method that finds aligned receptor pocket/probe protrusion pairs without explicit similarity computations. The RCF method enables the integration of systematic methods and local shape feature matching methods. To verify the efficiency and the accuracy of RCF, we integrated it with a systematic protein-protein docking approach (ATTRACT) based on a reduced protein representation. The test results show that the integrated docking approach is much faster. At the same time, it ranks the lowest ligand root-mean-square deviation (RMSD) (L_rms) solutions higher when docking enzyme-enzyme inhibitor complexes. Consequently, RCF not only enables much faster execution of systematic docking runs but also improves the qualities of docking predictions.     

Allometric variation among juvenile, adult male and female eastern bearded dragons Pogona barbata (Cuvier, 1829), with comments on the behavioural implications

The functional significance of allometric change in reptiles has received limited attention and the reason for such changes has been regarded as ‘obscure’. In this paper we report data on the Australian Pogona barbata, the eastern bearded dragon, from across their range and review changes in allometric growth among juveniles, and adult males and females and consider the functional relevance of these changes. There were significant differences in the population for mass, tail length, tail width, rear leg length and jaw length. These differences were consistent with differences required in locomotor performance and thus habitat use, together with access to different preferred dietary components.     

A fungal enzyme with the ability of aflatoxin B₁ conversion: purification and ESI-MS/MS identification

Armillariella tabescens, a Chinese edible and medicinal fungus, whose multienzyme exist ability of AFB₁-converting, and ADTZ (aflatoxin-detoxizyme) had previously purified from the A. tabescens multienzyme monitored by AFB₁ conversion_. However, the enzyme now confirmed an oxidase and renamed aflatoxin-oxidase (AFO). In this paper, AFO was purified by an economical and practical three-step procedure monitored by AFB₁ conversion. And ESI-MS/MS analysis was done for identification of AFO. The following database searching (Protein Blast on NCBI) results did not show any homologous oxidase protein, which implied that AFO was mostly a new oxidase differing from other reported aflatoxin-converting enzymes such as fungal laccase and horse radish peroxidase. HPTLC analysis of the purified AFO activity suggested that the enzyme reacted at the bisfuran ring of AFB₁ which was the key toxic structure. Therefore, all these investigations implied a new choice for biodegradation of aflatoxin in foods and feeds with the practical application of AFO.