DNA sequence plays a major role in determining nucleosome positions in yeast CUP1 chromatin.

The role of DNA sequence in determining nucleosome positions in vivo was investigated by comparing the positions adopted by nucleosomes reconstituted on a yeast plasmid in vitro using purified core histones with those in native chromatin containing the same DNA, described previously. Nucleosomes were reconstituted on a 2.5 kilobase pair DNA sequence containing the yeast TRP1ARS1 plasmid with CUP1 as an insert (TAC-DNA). Multiple, alternative, overlapping nucleosome positions were mapped on TAC-DNA. For the 58 positioned nucleosomes identified, the relative positioning strengths and the stabilities to salt and temperature were determined. These positions were, with a few exceptions, identical to those observed in native, remodeled TAC chromatin containing an activated CUP1 gene. Only some of these positions are utilized in native, unremodeled chromatin. These observations suggest that DNA sequence is likely to play a very important role in positioning nucleosomes in vivo. We suggest that events occurring in yeast CUP1 chromatin determine which positions are occupied in vivo and when they are occupied.     

ACE/ACE2 Ratio and MMP-9 Activity as Potential Biomarkers in Tuberculous Pleural Effusions

Objective: Pleural effusion is common problem, but the rapid and reliable diagnosis for specific pathogenic effusions are lacking. This study aimed to identify the diagnosis based on clinical variables to differentiate pleural tuberculous exudates from other pleural effusions. We also investigated the role of renin-angiotensin system (RAS) and matrix metalloproteinase (MMPs) in the pathogenesis of pleural exudates.Experimental design: The major components in RAS and extracellular matrix metabolism, including angiotensin converting enzyme (ACE), ACE2, MMP-2 and MMP-9 activities, were measured and compared in the patients with transudative (n = 45) and exudative (n = 80) effusions. The exudative effusions were come from the patients with tuberculosis (n = 20), pneumonia (n = 32), and adenocarcinoma (n = 28).Results: Increased ACE and equivalent ACE2 activities, resulting in a significantly increased ACE/ACE2 ratio in exudates, were detected compared to these values in transudates. MMP-9 activity in exudates was significantly higher than that in transudates. The significant correlation between ACE and ACE2 activity that was found in transudates was not found in exudates. Advanced analyses showed significantly increased ACE and MMP-9 activities, and decreased ACE2 activity in tuberculous pleural effusions compared with those in pneumonia and adenocarcinoma effusions. The results indicate that increased ACE and MMP-9 activities found in the exudates were mainly contributed from a higher level of both enzyme activities in the tuberculous pleural effusions.Conclusion: Interplay between ACE and ACE2, essential functions in the RAS, and abnormal regulation of MMP-9 probably play a pivotal role in the development of exudative effusions. Moreover, the ACE/ACE2 ratio combined with MMP-9 activity in pleural fluid may be potential biomarkers for diagnosing tuberculous pleurisy.     

MV-mimicking micelles loaded with PEG-serine-ACP nanoparticles to achieve biomimetic intra/extra fibrillar mineralization of collagen in vitro

Since their discovery, matrix vesicles (MVs) containing minerals have received considerable attention for their role in the mineralization of bone, dentin and calcified cartilage. Additionally, MVs' association with collagen fibrils, which serve as the scaffold for calcification in the organic matrix, has been repeatedly highlighted. The primary purpose of the present study was to establish a MVs–mimicking model (PEG-S-ACP/micelle) in vitro for studying the exact mechanism of MVs-mediated extra/intra fibrillar mineralization of collagen in vivo. In this study, high-concentration serine was used to stabilize the amorphous calcium phosphate (S-ACP), which was subsequently mixed with polyethylene glycol (PEG) to form PEG-S-ACP nanoparticles. The nanoparticles were loaded in the polysorbate 80 micelle through a micelle self-assembly process in an aqueous environment. This MVs–mimicking model is referred to as the PEG-S-ACP/micelle model. By adjusting the pH and surface tension of the PEG-S-ACP/micelle, two forms of minerals (crystalline mineral nodules and ACP nanoparticles) were released to achieve the extrafibrillar and intrafibrillar mineralization, respectively. This in vitro mineralization process reproduced the mineral nodules mediating in vivo extrafibrillar mineralization and provided key insights into a possible mechanism of biomineralization by which in vivo intrafibrillar mineralization could be induced by ACP nanoparticles released from MVs. Also, the PEG-S-ACP/micelle model provides a promising methodology to prepare mineralized collagen scaffolds for repairing bone defects in bone tissue engineering.     

Engineering lysine‐rich caleosins as carrier proteins to render biotin as a hapten on artificial oil bodies for antibody production

It has been demonstrated that caleosin alone is sufficient to stabilize artificial oil bodies. A series of recombinant caleosins, mutated with 3, 5, 8, 11, 13, 15, and 17 extra Lys residues and over‐expressed in Escherichia coli, were used as carrier proteins to render biotin as a hapten on the surface of artificial oil bodies for antibody production. Biotinylation levels of the recombinant caleosins were step‐wisely elevated as the number of extra Lys residues increased, and the biotinylated Lys residues were identified by mass spectrometric analysis. Polyclonal antibodies against biotin were successfully generated in rats injected with artificial oil bodies constituted with each of the biotinylated caleosins. Moreover, those generated via the biotinylated caleosins with eight or more extra Lys residues no longer recognized caleosin. It appears that engineered Lys‐rich caleosins are suitable carrier proteins for the production of antibodies against small molecules. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011     

Multiomic Metabolic Enrichment Network Analysis Reveals Metabolite–Protein Physical Interaction Subnetworks Altered in Cancer

Metabolism is recognized as an important driver of cancer progression and other complex diseases, but global metabolite profiling remains a challenge. Protein expression profiling is often a poor proxy since existing pathway enrichment models provide an incomplete mapping between the proteome and metabolism. To overcome these gaps, we introduce multiomic metabolic enrichment network analysis (MOMENTA), an integrative multiomic data analysis framework for more accurately deducing metabolic pathway changes from proteomics data alone in a gene set analysis context by leveraging protein interaction networks to extend annotated metabolic models. We apply MOMENTA to proteomic data from diverse cancer cell lines and human tumors to demonstrate its utility at revealing variation in metabolic pathway activity across cancer types, which we verify using independent metabolomics measurements. The novel metabolic networks we uncover in breast cancer and other tumors are linked to clinical outcomes, underscoring the pathophysiological relevance of the findings.