Function of yeast Rad52 protein as a mediator between replication protein A and the Rad51 recombinase

The RAD51 and RAD52 genes of Saccharomyces cerevisiae are key members of the RAD52 epistasis group required for genetic recombination and the repair of DNA double-stranded breaks. The RAD51 encoded product mediates the DNA strand exchange reaction. Efficient strand exchange is contingent upon the addition of the heterotrimeric single-stranded DNA binding factor replication protein A (RPA) after Rad51 has nucleated onto the single-stranded DNA. However, if the single-stranded DNA is incubated with Rad51 and RPA simultaneously to mimic what may be expected to occur in vivo, the efficiency of strand exchange decreases dramatically, revealing an inhibitory effect of RPA that is distinct from its stimulatory function. Interestingly, the inclusion of Rad52 protein, which has been purified in this study from yeast cells, restores the efficiency of strand exchange. Thus, Rad52 functions as a co-factor for the Rad51 recombinase, acting specifically to overcome the apparent competition by RPA for binding to single-stranded DNA.     

Requirement of cysteine-rich repeats of the Fas receptor for binding by the Fas ligand

The Fas receptor is a member of a family of cell death receptors, including tumor necrosis factor receptor I (TNFR I), death receptor 3 and 4 (DR3 and DR4), and cytopathic avian receptor 1 (CAR1). The Fas receptor is composed of several discrete domains, including three cysteine-rich domains (CRDs), a transmembrane domain, and an intracellular domain responsible for transmitting an apoptotic signal. While the mechanism of Fas-mediated cell death has become elucidated, the requirements for Fas ligand binding to the receptor have not been fully defined. Using a series of chimeric Fc-receptor fusion proteins between the human Fas receptor and TNFR I, each cysteine-rich domain of Fas was found to be required for interaction with the Fas ligand. Interestingly, TNFR I CRD1 could partially substitute for the Fas CRD1. The importance of this domain was underscored by the analysis of a Fas extracellular mutation (C66R), which resulted in a complete loss of ligand binding. This mutation was cloned from a human patient suffering from Canale-Smith syndrome, which is characterized by autoimmunity resembling that observed in the lpr and lprcg mice. The localization of essential ligand binding domains in the Fas receptor correlated exactly with the ability of the Fas receptor fusion proteins to prevent cell death mediated by the Fas ligand.     

A linear minimum mean square error multiuser receiver inRayleigh-fading channels

We generalize the multiuser (CDMA spread spectrum) communication systems to the fading environments. We first extend Verdu's (1986) conventional optimum receiver to Rayleigh-fading environments and then evaluate its performance. Having no knowledge of the received power at the receiving end, we therefore need an estimator to efficiently estimate the received signal strength of each user in fading environments. A linear minimum mean square error (LMMSE) unbiased estimator is proposed to attain this goal. By using the minimum mean square error (MMSE) Bayesian estimation, we further propose the LMMSE bit estimator for efficient demodulation. Its performance is close to the optimum multiuser receiver but with a much simpler polynomial complexity. To further reduce the complexity, we extend the LMMSE estimator to the sequential LMMSE estimator. In sequential estimation, we do not need to implement the matched filter banks and to perform the matrix inversion when estimating. In addition, it converges after approximately 2k iterations, where k is the number of users. With this fast convergence property and the simple structure, the sequential LMMSE estimator provides an attractive alternative to the implementation of a multiuser system     

Phase transformation of calcia-alumina-magnesia fibres produced by inviscid melt spinning

CaO-Al₂O₃-MgO (CAM) ceramic fibre produced via inviscid melt spinning (IMS) was investigated for phase transformation. Differential thermal analysis (DTA) on the as-spun CAM fibre gave two transformation peaks, one for exothermic peak at around 927°C and the other for endothermic one at around 1100°C. In order to identify each phase transformation x-ray diffraction (XRD) analysis was performed on the CAM fibres heat-treated to each phase transformation completion temperature. The exothermic peak was determined to represent crystallization of remaining amorphous phase in the as-spun CAM fibre. The endothermic peak was determined to correspond to transformation of non-equilibrium CaO · Al₂O₃ phase to equilibrium 3CaO· 5Al₂O₃ phase.     

Cryptanalysis of an involutional block cipher using cellular automata

In 2006, an involutional block cipher using cellular automata was proposed. A self-invertible CA-based structure allows for an efficient hardware implementation. This paper analyzes the insecurity of the cipher due to its conjugate property. The results of this study will make it possible to construct a decryption process without knowledge of the secret key.     

Microstructure of aragonite grown at an air–liquid interface

Aragonite particles dispersed in a bioresorbable polymer matrix are considered to be a good candidate for bone prosthesis materials. It is important to characterize the microstructure of synthetic aragonite used for biomedical applications, since the microstructure may influence its integration, resorption and replacement by bone. We studied late stages of aragonite growth, at an air–liquid interface, from a solution not doped with additives. Comparison was made between the types of synthetic aragonite microstructure and that of aragonite which is found in nature (mollusc shells, gallstones, Earth's crust). The microstructure of natural aragonite is unique to certain classes of living organisms and the understanding of its structure/function relationships may help to select the types of synthetic aragonite for specific biomedical applications. Three types of synthetic aragonite were observed based on grain size and grain morphology. This revised version was published online in November 2006 with corrections to the Cover Date.     

Identification of an interaction between the m-band protein skelemin and beta-integrin subunits. Colocalization of a skelemin-like protein with beta1- and beta3-integrins in non-muscle cells

Signaling across integrins is regulated by interaction of these receptors with cytoskeletal proteins and signaling molecules. To identify molecules interacting with the cytoplasmic domain of the beta3-integrin subunit (glycoprotein IIIa), a placental cDNA library was screened in the yeast two-hybrid system. Two identical clones coding for a 96-amino acid sequence were identified. This sequence was 100% identical to a sequence in skelemin, a protein identified previously in skeletal muscle. Skelemin is a member of a superfamily of cytoskeletal proteins that contain fibronectin-type III-like motifs and immunoglobulin C2-like motifs and that regulate the organization of myosin filaments in muscle. The amino acid residues in the isolated clones encompassed C2 motifs 4 and 5 of skelemin. A recombinant skelemin protein consisting of C2 motifs 3-7 interacted with beta1- and beta3-integrin cytoplasmic domains expressed as glutathione S-transferase (GST) fusion proteins, but not with GST-beta2-integrin cytoplasmic tail or GST alone. The skelemin-binding region was in the membrane proximal cytoplasmic domains of the integrins. Full-length skelemin interacted with integrin in intact cells as demonstrated by the colocalization of hemagglutinin-tagged skelemin in Chinese hamster ovary (CHO) cells containing alphaIIbbeta3-integrin and by the finding that microinjection of C2 motif 4 of skelemin into C2C12 mouse myoblast cells caused spread cells to round up. A skelemin-like protein was detected in CHO cells, endothelial cells, and platelets, and this protein colocalized with beta1- and beta3-integrins in CHO cells. This study suggests the presence of a skelemin-like protein in non-muscle cells and provides evidence that it may be involved in linking integrins to the cytoskeleton.     

Identification of an opioid kappa receptor subtype-selective N-substituent for (+)-(3R,4R)-dimethyl-4-(3-hydroxyphenyl)piperidine

A three-component library of compounds was prepared in parallel using multiple simultaneous solution-phase synthetic methodology. The compounds were biased toward opioid receptor antagonist activity by incorporating (+)-(3R,4R)-dimethyl-4-(3-hydroxyphenyl)piperidine (a potent, nonselective opioid pure antagonist) as one of the monomers. The other two monomers, which included N-substituted or unsubstituted Boc-protected amino acids and a range of substituted aryl carboxylic acids, were selected to add chemical diversity. Screening of these compounds in competitive binding experiments with the kappa opioid receptor selective ligand [3H]U69,593 led to the discovery of a novel kappa opioid receptor selective ligand, N-?(2'S)-[3-(4-hydroxyphenyl)propanamido]-3'-methylbutyl?-(3R, 4R)-dimethyl-4-(3-hydroxyphenyl)piperidine (8, RTI-5989-29). Additional structure-activity relationship studies suggested that 8 possesses lipophilic and hydrogen-bonding sites that are important to its opioid receptor potency and selectivity. These sites appear to exist predominantly within the kappa receptor since the selectivity arises from a 530-fold loss of affinity of 8 for the mu receptor and an 18-fold increase in affinity for the kappa receptor relative to the mu-selective ligand, (+)-N-[trans-4-phenyl-2-butenyl]-(3R, 4R)-dimethyl-4-(3-hydroxyphenyl)piperidine (5a). The degree of selectivity observed in the radioligand binding experiments was not observed in the functional assay. According to its ability to inhibit agonist stimulated binding of [35S]GTPgammaS at all three opioid receptors, compound 8 behaves as a mu/kappa opioid receptor pure antagonist with negligible affinity for the delta receptor.     

RANTES-induced T cell activation correlates with CD3 expression

The chemokine RANTES induces a unique biphasic cytoplasmic Ca2+ signal in T cells. The first phase of this signal, similar to that of other chemokines, is G-protein mediated and chemotaxis associated. The second phase of this signal, unique to RANTES and evident at concentrations greater than 100 nM, is tyrosine kinase linked and results in a spectrum of responses similar to those seen with antigenic stimulation of T cells. We show here that certain jurkat T cells responded to RANTES solely through this latter pathway. A direct correlation between the RANTES-induced second phase response and CD3 expression was demonstrated in these cells. Sorting the Jurkat cells into CD3(high) and CD3(low) populations revealed that only the CD3(high) cells were responsive to RANTES. Furthermore, stimulation of these Jurkat cells with anti-CD3 mAb significantly depresses their subsequent response to RANTES. While a RANTES-specific chemokine receptor is expressed at a low level on these Jurkat cells, the RANTES-induced activation is dependent on the presence of the TCR. Thus, stimulation through TCR may partially account for RANTES' unique pattern of signaling in T cells.