Radiation inactivation analysis of H+-pyrophosphatase from submitochondrial particles of etiolated mung bean seedlings

AbstractRadiation inactivation analysis was employed to determine the functional masses of enzymatic activity and proton translocation of H+-pyrophosphatase from submitochondrial particles of etiolated mung bean seedlings. The activities of H+-pyrophosphatase decayed as a simple exponential function with respect to radiation dosage. D37 values of 6.9±0.3 and 7.5±0.5 Mrad were obtained for pyrophosphate hydrolysis and its associated proton translocation, yielding molecular masses of 170±7 and 156±11 kDa, respectively. In the presence of valinomycin and 50 mM KCl, the functional size of H+-pyrophosphatase of tonoplast was decreased, while that of submitochondrial particles remained the same, indicating that they are two distinct types of proton pump using PPi as their energy source

Profiling time course expression of virus genes---an illustration of Bayesian inference under shape restrictions

There have been several studies of the genome-wide temporal transcriptional program of viruses, based on microarray experiments, which are generally useful in the construction of gene regulation network. It seems that biological interpretations in these studies are directly based on the normalized data and some crude statistics, which provide rough estimates of limited features of the profile and may incur biases. This paper introduces a hierarchical Bayesian shape restricted regression method for making inference on the time course expression of virus genes. Estimates of many salient features of the expression profile like onset time, inflection point, maximum value, time to maximum value, area under curve, etc. can be obtained immediately by this method. Applying this method to a baculovirus microarray time course expression data set, we indicate that many biological questions can be formulated quantitatively and we are able to offer insights into the baculovirus biology.Comment: Published in at http://dx.doi.org/10.1214/09-AOAS258 the Annals of Applied Statistics (http://www.imstat.org/aoas/) by the Institute of Mathematical Statistics (http://www.imstat.org

Peroxisome Proliferator–Activated Receptor γ Level Contributes to Structural Integrity and Component Production of Elastic Fibers in the AortaNovelty and Significance

Loss of integrity and massive disruption of elastic fibers are key features of abdominal aortic aneurysm (AAA). Peroxisome proliferator-activated receptor γ (PPARγ) has been shown to attenuate AAA through inhibition of inflammation and proteolytic degradation. However, its involvement in elastogenesis during AAA remains unclear. PPARγ was highly expressed in human AAA within all vascular cells, including inflammatory cells and fibroblasts. In the aortas of transgenic mice expressing PPARγ at 25% normal levels (PpargC/− mice), we observed the fragmentation of elastic fibers and reduced expression of vital elastic fiber components of elastin and fibulin-5. These were not observed in mice with 50% normal PPARγ expression (Pparg+/− mice). Infusion of a moderate dose of angiotensin II (AngII) (500 ng/kg/min) did not induce AAA but Pparg+/− aorta developed flattened elastic lamellae, while PpargC/− aorta showed severe destruction of elastic fibers. After infusion of AngII at 1000 ng/kg/min, 73% of PpargC/− mice developed atypical suprarenal aortic aneurysms: superior mesenteric arteries were dilated with extensive collagen deposition in adventitia and infiltrations of inflammatory cells. Although matrix metalloproteinase inhibition by doxycycline somewhat attenuated the dilation of aneurysm, it did not reduce the incidence nor elastic lamella deterioration in AngII-infused PpargC/− mice. Furthermore, PPARγ antagonism down-regulated elastin and fibulin-5 in fibroblasts, but not in vascular smooth muscle cells. Chromatin immunoprecipitation assay demonstrated PPARγ binding in the genomic sequence of fibulin-5 in fibroblasts. Our results underscore the importance of PPARγ in AAA development though orchestrating proper elastogenesis and preserving elastic fiber integrity

Development of a Surface Plasmon Resonance Biosensor for Real-Time Detection of Osteogenic Differentiation in Live Mesenchymal Stem Cells

Surface plasmon resonance (SPR) biosensors have been recognized as a useful tool and widely used for real-time dynamic analysis of molecular binding affinity because of its high sensitivity to the change of the refractive index of tested objects. The conventional methods in molecular biology to evaluate cell differentiation require cell lysis or fixation, which make investigation in live cells difficult. In addition, a certain amount of cells are needed in order to obtain adequate protein or messenger ribonucleic acid for various assays. To overcome this limitation, we developed a unique SPR-based biosensing apparatus for real-time detection of cell differentiation in live cells according to the differences of optical properties of the cell surface caused by specific antigen-antibody binding. In this study, we reported the application of this SPR-based system to evaluate the osteogenic differentiation of mesenchymal stem cells (MSCs). OB-cadherin expression, which is up-regulated during osteogenic differentiation, was targeted under our SPR system by conjugating antibodies against OB-cadherin on the surface of the object. A linear relationship between the duration of osteogenic induction and the difference in refractive angle shift with very high correlation coefficient was observed. To sum up, the SPR system and the protocol reported in this study can rapidly and accurately define osteogenic maturation of MSCs in a live cell and label-free manner with no need of cell breakage. This SPR biosensor will facilitate future advances in a vast array of fields in biomedical research and medical diagnosis

Recovery of heat shock-triggered released apoplastic Ca2+ accompanied by pectin methylesterase activity is required for thermotolerance in soybean seedlings

Synthesis of heat shock proteins (HSPs) in response to heat shock (HS) is essential for thermotolerance. The effect of a Ca2+ chelator, EGTA, was investigated before a lethal HS treatment in soybean (Glycine max) seedlings with acquired thermotolerance induced by preheating. Such seedlings became non-thermotolerant with EGTA treatment. The addition of Ca2+, Sr2+ or Ba2+ to the EGTA-treated samples rescued the seedlings from death by preventing the increased cellular leakage of electrolytes, amino acids, and sugars caused by EGTA. It was confirmed that EGTA did not affect HSP accumulation and physiological functions but interfered with the recovery of HS-released Ca2+ concentration which was required for thermotolerance. Pectin methylesterase (PME, EC 3.1.1.11), a cell wall remodelling enzyme, was activated in response to HS, and its elevated activity caused an increased level of demethylesterified pectin which was related to the recovery of the HS-released Ca2+ concentration. Thus, the recovery of HS-released Ca2+ in Ca2+-pectate reconstitution through PME activity is required for cell wall remodelling during HS in soybean which, in turn, retains plasma membrane integrity and co-ordinates with HSPs to confer thermotolerance

MRE11 promotes oral cancer progression through RUNX2/CXCR4/AKT/FOXA2 signaling in a nuclease-independent manner

MRE11, the nuclease component of RAD50/MRE11/NBS1 DNA repair complex which is essential for repair of DNA double-strand-breaks in normal cells, has recently garnered attention as a critical factor in solid tumor development. Herein we report the crucial role of MRE11 in oral cancer progression in a nuclease-independent manner and delineate its key downstream effectors including CXCR4. MRE11 expression in oral cancer samples was positively associated with tumor size, cancer stage and lymph node metastasis, and was predictive of poorer patient survival and radiotherapy resistance. MRE11 promoted cell proliferation/migration/invasion in a nuclease-independent manner but enhanced radioresistance via a nuclease-dependent pathway. The nuclease independent promotion of EMT and metastasis was mediated by RUNX2, CXCR4, AKT, and FOXA2, while CXCR4 neutralizing antibody mitigated these effects in vitro and in vivo. Collectively, MRE11 may serve as a crucial prognostic factor and therapeutic target in oral cancer, displaying dual nuclease dependent and independent roles that permit separate targeting of tumor vulnerabilities in oral cancer treatment