Actin-related proteins (Arps) and conventional actin are enigmatic components of many chromatin-remodeling enzyme complexes. The yeast INO80 ATP-dependent chromatin-remodeling complex contains stoichiometric amounts of Arp4, Arp5, Arp8, and actin. Here we have revealed functions of Arp5 and Arp8 by analysis of mutants. arp5 Delta and arp8 Delta mutants display an ino80 Delta phenotype. Purification of INO80 complexes from arp5 Delta and arp8 Delta cells shows that protein complexes remain intact but are compromised for INO80 ATPase activity, DNA binding, and nucleosome mobilization. The INO80 (arp8 Delta) complex is strikingly deficient, not only for the Arp8 subunit, but also for Arp4 and actin, suggesting an ordered assembly of Arps. Binding of Arp8 to the INO80 complex requires an N-terminal region of Ino80 adjacent to the conserved ATPase domain. GST-Arp8 binds preferentially to histones H3 and H4 in vitro, suggesting a histone chaperone function. These findings show direct involvement of Arps in the chromatin-remodeling process.

To enhance the osteogenic activity of BMP, combination BMP2 and BMP7 gene transfer was performed. This approach led to a significant increase in osteoblastic differentiation of mesenchymal precursors compared with single BMP gene transfer in vitro. When tested in 78 rats, combination gene transfer enhanced mechanically stable spine fusion and bone formation rate versus single BMP gene transfer.Introduction: Although clinical bone morphogenetic protein (BMP) therapy is effective, required doses are very high. Previous studies have suggested that the co-expression of two different BMP genes can result in the production of heterodimeric BMPs that may be more potent than homodimers. In this study, combined BMP2 and BMP7 gene transfer was performed to test whether this approach improves osteoblastic differentiation and bone formation compared with single BMP gene transfer. Materials and Methods: A producer cell (A549) was co-transfected with adenovirus vectors encoding BMP2 (AdBMP2) and BMP7 (AdBMP7) or, as controls, each vector alone, AdNull (with no transgene) or no virus. Supernatants were compared for their ability to stimulate osteoblastic differentiation of C2C12 myoblasts and MC3T3-E1 pre-osteoblasts. In a rat posterolateral spine fusion model, co-administration of AdBMP2 and AdBMP7 was compared with treatment with each vector alone, AdNull or no virus in 78 rats. The spines were assessed 8 weeks after surgery for radiographic and mechanical fusion, bone formation, and mineralization. Results: BMP2 and BMP7 were co-precipitated from supernatants of cells co-transfected with AdBMP2 and AdBMP7, indicating the presence of BMP2/7 heterodimer. Supernatants of co-transfected cells containing relatively low doses (7-140 ng/ml) of BMPs induced osteocalcin expression and alkaline phosphatase activity in both C2C12 and MC3T3-E1 cells, that were up to 6-and 40-fold higher, respectively, than levels induced by maximal doses (200 -1000 ng/ml) of either BMP2 or BMP7 alone. In the spine fusion model, co-administration of AdBMP2 and AdBMP7 resulted in a significantly greater number of mechanically stable fusions and also 2-fold higher mineralization rate and bone volume in the fusion mass versus single BMP gene transfer (p Ͻ 0.02, all comparisons). Conclusion: Combined BMP2 and BMP7 gene transfer is significantly more effective in inducing osteoblastic differentiation and spine fusion than individual BMP gene transfer.

BackgroundPulmonary actinomycosis is a chronic pulmonary infection caused by Actinomyces. Both improving oral hygiene and early application of antibiotics to the case of suspicious pulmonary infections result in changes in incidences and presentations of pulmonary actinomycosis. However, there are little reports dealt with the recent clinical characteristics of pulmonary actinomycosis. This study aimed to review the characteristics of pulmonary actinomycosis occurred during the first decade of 21st century.MethodsThis retrospective study was performed on 94 subjects with pulmonary actinomycosis diagnosed pathologically from January 2000 to December 2010 in 13 hospitals in Korea.ResultsThe data of the study showed that pulmonary actinomycosis occurs frequently in middle to old-aged males (mean age; 57.7 years old) and that the most common symptoms are cough, hemoptysis, and sputum production. Various radiologic features such as the consolidation with central low attenuation (74.5%) and no regional predominance were also observed. Most of patients recovered completely with medical and/or surgical treatment, reaching approximately 98% cure rate.ConclusionsThe results demonstrate that pulmonary actinomycosis is one of the cautious pulmonary diseases. More importantly, in cases of persistent hemoptysis or for differential diagnosis from lung malignancy, our data have revealed that surgical resection appears to be a useful intervention and that radiologic diagnosis may not provide decisive information. These findings indicate that it is important for the clinicians to include pulmonary actinomycosis as one of differential diagnoses for refractory pulmonary abnormal lesions to the current usual management.

Stress hormones have been implicated in both tumor initiation and progression. Human telomerase reverse transcriptase (hTERT) is overexpressed in cancer cells and associated with malignant tumor progression and poor outcome. We thus sought to determine whether the stress hormone norepinephrine (NE) could induce hTERT expression and subsequently ovarian cancer progression. Unexpectedly, NE induced hTERT transcript and protein expression, and subsequently ovarian cancer cell invasion. Pharmacologic inhibition of β2-adrenergic receptor 2 and protein kinase A, as well as silencing of hypoxia-inducible factor-1α and c-Myc expression, profoundly attenuated NE-induced hTERT expression. Strikingly, stimulation of the cells with NE or ectopic expression of hTERT induced expression of Slug, ovarian cancer cell epithelial-mesenchymal transition (EMT) and invasion. Silencing of hTERT expression abrogated NE-induced ovarian cancer cell invasion, EMT and Slug expression. In addition, silencing of Slug expression significantly inhibited NE- and hTERT-induced ovarian cancer cell EMT and invasion. Moreover, continuous exposure to NE was sufficient to enhance in vivo hTERT expression and metastasis of ovarian cancer cells to the lung. Finally, we provide evidence that hTERT links Src to Slug expression in NE-induced ovarian cancer EMT and metastasis. We thus demonstrate a novel role of hTERT in stress hormone-induced ovarian cancer aggressiveness through inducing Slug, providing novel biomarkers and potential therapeutic targets for ovarian cancer.

WHO Grades II and III meningiomas occur far less frequently in the cranial base and spinal locations. Tumors arising from these locations may have different mechanisms of tumorigenesis and/or progression compared with meningiomas arising from other (non-cranial base) regions.

These findings suggest that DNA methylation may be important for downregulation of CFTR gene expression in lung cancer. Promoter hypermethylation of the CFTR gene may be an important prognostic factor in younger patients with NSCLC.

The adsorption of membrane-associated protein cytochrome c to anionic lipid bilayers of dioleoyl phosphatidylglycerol was studied in low ionic strength physiological buffer using atomic force microscopy. The bilayers were supported on polylysinated mica. The formation of stable, single lipid bilayers was confirmed by imaging and force spectroscopy. Upon addition of low concentrations of cytochrome c, protein molecules were not topographically visible on the lipid bilayer-buffer interface. However, the forces required to punch through the bilayer by indentation using the atomic force microscopy probe were significantly lower after protein adsorption, which suggest that the protein inserts into the bilayer. Moreover, the apparent thickness of the bilayer remained unchanged after cytochrome c adsorption. Yet, mass spectroscopy and visible light absorption spectroscopy confirmed the presence of cytochrome c in the lipid bilayers. These results suggest that 1), cytochrome c inserts into the bilayer and resides in its hydrophobic core; 2), cytochrome c insertion changes the mechanical properties of the bilayer significantly; and 3), bilayer force spectroscopy may be a useful tool in investigating lipid-protein interactions.

In bacteria, the contribution of global nucleoid organization in determining cellular transcription programs is unclear. Using a mutant form of the most abundant nucleoid-associated protein HU, HU␣ E38K,V42L , we previously showed that nucleoid remodeling by the mutant protein re-organizes the global transcription pattern. Here, we demonstrate that, unlike the dimeric wild-type HU, HU␣ E38K,V42L is an octamer and wraps DNA around its surface. The formation of wrapped nucleoprotein complexes by HU␣ E38K,V42L leads to a high degree of DNA condensation. The DNA wrapping is right-handed, which restrains positive supercoils. In vivo, HU␣ E38K,V42L shows altered association and distribution patterns with the genetic loci whose transcription are differentially affected in the mutant strain.Chromatin is defined as the complex conglomeration of chromosomal DNA and specific DNA-binding proteins. Its organization is dynamically attuned to various changes in transcriptional requirements in response to exogenous and endogenous cues. In eukaryotes, the basic unit of chromatin is the nucleosome, which consists of DNA wrapped in left-handed turns around a histone octamer (1). In prokaryotes, on the other hand, it is commonly believed that the chromosome (nucleoid) does not have a defined structure. Nevertheless, bacterial nucleoids are efficiently compacted by a disparate group of histone-like DNA-binding proteins with low molecular mass and high electrostatic charge. HU is one of the most conserved and abundant (50,000 dimers per cell during logarithmic phase) nucleoid-associated proteins identified in eubacteria (2). In most bacteria, HU exists as a 18-kDa homodimer. Only in Enterobacteriaceae, including Escherichia coli, HU is a heterodimer of two similar subunits, HU␣ and HU␤. HU has traditionally been accepted as the archetypal bacterial counterpart of eukaryotic histones and evolutionarily linked to histone H1 (3). HU is a nonspecific DNA-binding protein, which plays an architectural role in bending DNA (4, 9) and constraining negative supercoils (5). It also participates in specific control functions in DNA transactions like replication, transcription, recombination, and DNA repair (6 -10). Earlier observations of compact nucleosome-like structures in DNA induced by HU binding (11,12) were recently challenged by studies that suggested that HU might in fact counteract DNA compaction by antagonizing other DNA-condensing proteins like H-NS (13). There has also been conflicting reports about the exact contribution of HU toward global chromosomal superhelicity (14). Thus, despite HU being one of the most abundant nucleoidassociated proteins, the exact molecular mechanisms of HU action with regard to its role in chromosome architecture and function remain largely obscure and controversial.We have recently identified a gain-of-function mutant form of HU␣, HU␣ E38K,V42L , which caused a reconfiguration of the E. coli nucleoid from a loosely packed, dispersed structure into a densely condensed, globular conformation (15). The mut...