Biofilm characteristics and evaluation of the sanitation procedures of thermophilic Aeribacillus pallidus E334 biofilms

The ability of Aeribacillus pallidus E334 to produce pellicle and form a biofilm was studied. Optimal biofilm formation occurred at 60 °C, pH 7.5 and 1.5% NaCl. Extra polymeric substances (EPS) were composed of proteins and eDNA (21.4 kb). E334 formed biofilm on many surfaces, but mostly preferred polypropylene and glass. Using CLSM analysis, the network-like structure of the EPS was observed. The A. pallidus biofilm had a novel eDNA content. DNaseI susceptibility (86.8% removal) of eDNA revealed its importance in mature biofilms, but the purified eDNA was resistant to DNaseI, probably due to its extended folding outside the matrix. Among 15 cleaning agents, biofilms could be removed with alkaline protease and sodium dodecyl sulphate (SDS). The removal of cells from polypropylene and biomass on glass was achieved with combined SDS/alkaline protease treatment. Strong A. pallidus biofilms could cause risks for industrial processes and abiotic surfaces must be taken into consideration in terms of sanitation procedures.     

The unique region of surrogate light chain component lambda5 is a heavy chain-specific regulator of precursor B cell receptor signaling

Signals transduced by precursor-BCRs (pre-BCRs) composed of Ig mu heavy chains (HCs) and the surrogate L chain components lambda5 and VpreB are critical for B cell development. A conserved unique region (UR) of lambda5 was shown to activate pre-BCR complexes in transformed cells and to engage putative ligands, but its contribution to pre-B cell development is not known. It is also not clear why the lambda-like sequences in lambda5 are used to select HCs that will associate mainly with kappa L chains. In this study, we show that, in transformed and primary mouse B cell progenitors, receptors containing full-length HCs and lacking the lambda5UR were expressed at higher surface levels, but exhibited reduced activity compared with normal pre-BCRs in supporting developmental changes that accompany the progenitor to pre-B cell transition in primary cell culture systems and in the bone marrow in vivo. In contrast, deletion of the lambda5UR did not change net signaling output by the Dmu-pre-BCR, a developmentally defective receptor that exhibited impaired activity in the primary cell culture system. Moreover, the lambda-like sequences in lambda5 were more accommodating than kappa in supporting surface expression and signaling by the different HCs. These results show that the lambda5UR is important, although not essential, for surrogate L chain-dependent receptor signaling in primary cells, and furthermore may help allow discrimination of signaling competency between normal and Dmu-pre-BCRs. That the lambda-like portion of lambda5 in the absence of the UR was nondiscriminatory suggests that the lambda5UR focuses pre-BCR-dependent selection on the HC V region.     

3D-QSAR analysis on benzazole derivatives as eukaryotic topoisomerase II inhibitors by using comparative molecular field analysis method

Selective topoisomerase II inhibitors have created a great deal of interest in recent years for the design of new antitumoral compounds. 3D-QSAR analysis has been performed on a series of previously synthesized benzoxazole, benzimidazole, and oxazolo(4,5-b)pyridine derivatives, which are screened as eukaryotic topoisomerase II inhibitors, using comparative molecular field analysis (CoMFA) with partial least squares fit to predict the steric and electrostatic molecular field interactions for the activity. The CoMFA study was carried out using a training set of 16 compounds. The predictive ability of the model was assessed using a test set of 7 compounds. The analyzed 3D-QSAR CoMFA model has demonstrated a good fit, having r(2) value of 0.997 and cross-validated coefficient q(2) value as 0.435 for the model. The obtained model reveals that the electronegatively charged substituents such as NO(2) or COOCH(3) group on position R and/or R(1) at the heterocyclic ring system and positively charged atom and/or atom groups located between the benzazole moiety and 2-substituted phenyl ring as a bridge element improve the activity. On the other hand, a bulky substituent, such as methoxy group, attached to the ortho position of 2-phenyl-5-nitro-benzoxazole (1) enhances the activity similar to compound 13, which is both a meta and para substituent of the phenyl group attached to the 2-position of benzimidazole ring system, fit into the favored steric region to improve the activity.     

A new composite facial and scalp transplantation model in rats

There are limited sources of autogenous tissue available for reconstruction of severe facial and scalp deformities caused by extensive tumor ablation, burns, or trauma. Allografts from cadaveric sources may serve as a reconstructive alternative. However, technical and immunological aspects of harvesting and transplanting face and scalp flaps limit the routine use of such procedures. For evaluation of the feasibility of composite-tissue reconstruction, an experimental model of composite face/scalp flap transplantation in rats was designed. Technical aspects of the model, survival rates, and the complications encountered during development of the model are presented. A total of 64 animals, in three experimental groups, were studied. In group I, the anatomical study group (n = 6), the anatomical features of the face and scalp region in rats were explored. Groups II and III were the transplantation groups. Isograft transplantations were performed between identical Lewis rats (RT11 to RT11), and allografts were transplanted, across major histocompatibility complex barriers, between Lewis-Brown Norway rats (RT1l/n) and Lewis rats (RT11). In group II (the control group, n = 8), transplantation of nonvascularized composite face/scalp isografts and allografts was performed. In group III (the transplantation group, n = 50), vascularized face/scalp isografts (n = 36) and allografts (n = 14) were transplanted. Complications included partial or total flap necrosis, death attributable to food aspiration, and poor general condition. To prevent acute and chronic allograft rejection, cyclosporine A (16 mg/kg per day) therapy was initiated 24 hours after transplantation; the dose was tapered to 2 mg/kg per day within 4 weeks and was maintained at that level thereafter. Long-term survival (>170 days) was achieved, without any signs of rejection, with low-dose (2 mg/kg per day) cyclosporine A therapy. This is the first report documenting successful composite face/scalp flap transplantation in the rat model.     

Latest developments in experimental and computational approaches to characterize protein–lipid interactions

Understanding the functional roles of all the molecules in cells is an ultimate goal of modern biology. An important facet is to understand the functional contributions from intermolecular interactions, both within a class of molecules (e.g. protein–protein) or between classes (e.g. protein‐DNA). While the technologies for analyzing protein–protein and protein–DNA interactions are well established, the field of protein–lipid interactions is still relatively nascent. Here, we review the current status of the experimental and computational approaches for detecting and analyzing protein–lipid interactions. Experimental technologies fall into two principal categories, namely solution‐based and array‐based methods. Computational methods include large–scale data‐driven analyses and predictions/dynamic simulations based on prior knowledge of experimentally identified interactions. Advances in the experimental technologies have led to improved computational analyses and vice versa, thereby furthering our understanding of protein–lipid interactions and their importance in biological systems.     

Mutational signatures of de-differentiation in functional non-coding regions of melanoma genomes

Much emphasis has been placed on the identification, functional characterization, and therapeutic potential of somatic variants in tumor genomes. However, the majority of somatic variants lie outside coding regions and their role in cancer progression remains to be determined. In order to establish a system to test the functional importance of non-coding somatic variants in cancer, we created a low-passage cell culture of a metastatic melanoma tumor sample. As a foundation for interpreting functional assays, we performed whole-genome sequencing and analysis of this cell culture, the metastatic tumor from which it was derived, and the patient-matched normal genomes. When comparing somatic mutations identified in the cell culture and tissue genomes, we observe concordance at the majority of single nucleotide variants, whereas copy number changes are more variable. To understand the functional impact of non-coding somatic variation, we leveraged functional data generated by the ENCODE Project Consortium. We analyzed regulatory regions derived from multiple different cell types and found that melanocyte-specific regions are among the most depleted for somatic mutation accumulation. Significant depletion in other cell types suggests the metastatic melanoma cells de-differentiated to a more basal regulatory state. Experimental identification of genome-wide regulatory sites in two different melanoma samples supports this observation. Together, these results show that mutation accumulation in metastatic melanoma is nonrandom across the genome and that a de-differentiated regulatory architecture is common among different samples. Our findings enable identification of the underlying genetic components of melanoma and define the differences between a tissue-derived tumor sample and the cell culture created from it. Such information helps establish a broader mechanistic understanding of the linkage between non-coding genomic variations and the cellular evolution of cancer.     

Proteomic identification of erythrocyte membrane protein deficiency in hereditary spherocytosis

Hereditary spherocytosis (HS) is the most common congenital hemolytic anemia in Caucasians, with an estimated prevalence ranging from 1:2000 to 1:5000. The molecular defect in one of the erythrocytes (RBC) membrane proteins underlying HS like; spectrin-α, spectrin-β, ankyrin, band 3 and protein 4.2 that lead to membrane destabilization and vesiculation, may change the RBCs into denser and more rigid cells (spherocytes), which are removed by the spleen, leading to the development of hemolytic anemia. It is classified as mild, moderate and severe, according to the degree of the hemolytic anemia and the associated symptoms. Two-dimensional gel electrophoresis (2-DE) is potentially valuable method for studying heritable disorders as HS that involve membrane proteins. This separation technique of proteins based upon two biophysically unrelated parameters; molecular weight and charge, is a good option in clinical proteomics in terms of ability to separate complex mixtures, display post-translational modifications and changes after phosphorylation. In this study, we have used contemporary methods with some modifications for the solubilisation, separation and identification of erythrocyte membrane proteins in normal and in HS RBCs. Spectrin alpha and beta chain, ankyrin and band 3 proteins expression differences were found with PDQuest software 8.0.1. and peptide mass fingerprinting (PMF) analysis performed for identification of proteins in this study.