The In Vitro Studies On Phage Mu Transposase

Bacteriophage Mu is one of the model systems to study DNA transposition. The availability of an in vitro soluble system greatly facilitates the dissection of the transposition mechanisms at the molecular levels. Several aspects of the in vitro mini-Mu DNA transposition have been dealt with in this thesis.;First, the flanking host DNA sequences on the strand cleavage step mediated by Mu transposase (Mu A protein) was investigated. We found that certain flanking host sequences could inhibit the strand cleavage step without affecting the earlier synapsis step. Furthermore, this cleavage defect could be overcome by the Mu B protein in the presence of ATP.;Second, the role of the 10 kDa C-terminal domain (domain III) of Mu transposase was studied. We showed that the cloned domain III was still functional in interacting with the Mu B protein in the absence of the 65 kDa N-terminal domain of Mu transposase. Deletion analysis revealed that the last 36 residues at the C-terminus of Mu transposase were involved in interacting with the Mu B protein. An intact C-terminus was required for efficient interactions between the Mu A and Mu B proteins.;Third, we mapped a novel non-specific DNA binding and nuclease activity in a 26 residue region (aa575-600) at N-terminus of domain III of Mu transposase. We showed that this region was required in both early synapsis step and the subsequent strand cleavage step. We argue that this 26 residue region might contact the Mu-host junctions in the transpososomes. Complementation studies further suggest that the active sites for the strand cleavage activity of Mu transposase are made up of amino acids in this 26 residue region of domain III on one transposase monomer and the conserved acidic residues (D, D35E) of domain II on a separate transposase monomer

A New Scheme and Microstructural Model for 3D Full 5-directional Braided Composites

AbstractThree-dimensional(3D) braided composites are a kind of advanced ones and are used in the aeronautical and astronautical fields more widely. The advantages, usages, shortages and disadvantages of 3D braided composites are analyzed, and the possible approach of improving the properties of the materials is presented, that is, a new type of 3D full 5-directional braided composites is developed. The methods of making this type of preform are proposed. It is pointed out that the four-step braiding which is the most possible to realize industrialized production almost has no effect on the composites'properties. By analyzing the simulation model, the advantages of the material compared with the 3D 4-di- rectional and 5-directional materials are presented. Finally, a microstructural model is analyzed to lay the foundation for the future theoretical analysis of these composites

Loss of STAT1 in Bone Marrow-Derived Cells Accelerates Skeletal Muscle Regeneration

BACKGROUND: Skeletal muscle regeneration is a complex process which is not yet completely understood. Evidence suggested that the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway may have a role in myogenesis. In this study, we aim to explore the possible role of STAT1 in muscle regeneration. METHODS: Wild-type and STAT1 knockout mice were used in this study. Tibialis anterior muscle injury was conducted by cardiotoxin (CTX) injection. Bone marrow transplantation and glucocorticoid treatment were performed to manipulate the immune system of the mice. RESULTS: Muscle regeneration was accelerated in STAT1-/- mice after CTX injury. Bone marrow transplantation experiments showed that the regeneration process relied on the type of donor mice rather than on recipient mice. Levels of pro-inflammatory cytokines, TNFα and IL-1β, were significantly higher in STAT1-/- mice at 1 day and/or 2 days post-injury, while levels of anti-inflammatory cytokine, IL-10, were lower in STAT1-/- mice at 2 days and 3 days post-injury. Levels of IGF-1 were significantly higher in the STAT1-/- mice at 1 day and 2 days post-injury. Furthermore, the muscle regeneration process was inhibited in glucocorticoid-treated mice. CONCLUSIONS: Loss of STAT1 in bone marrow-derived cells accelerates skeletal muscle regeneration

Autophagy Protects the Blood-Brain Barrier Through Regulating the Dynamic of Claudin-5 in Short-Term Starvation

The blood-brain barrier (BBB) is essential for the exchange of nutrient and ions to maintain the homeostasis of central nervous system (CNS). BBB dysfunction is commonly associated with the disruption of endothelial tight junctions and excess permeability, which results in various CNS diseases. Therefore, maintaining the structural integrity and proper function of the BBB is essential for the homeostasis and physiological function of the CNS. Here, we showed that serum starvation disrupted the function of endothelial barrier as evidenced by decreased trans-endothelial electrical resistance, increased permeability, and redistribution of tight junction proteins such as Claudin-5 (Cldn5). Further analyses revealed that autophagy was activated and protected the integrity of endothelial barrier by scavenging ROS and inhibiting the redistribution of Cldn5 under starvation, as evidenced by accumulation of autophagic vacuoles and increased expression of LC3II/I, ATG5 and LAMP1. In addition, autophagosome was observed to package and eliminate the aggregated Cldn5 in cytosol as detected by immunoelectron microscopy (IEM) and stimulated emission depletion (STED) microscope. Moreover, Akt-mTOR-p70S6K pathway was found to be involved in the protective autophagy induced by starvation. Our data demonstrated that autophagy played an essential role in maintaining the integrity of endothelial barrier by regulating the localization of Cldn5 under starvation