A Multi-Scale Approach for Phase Field Modeling of Ultra-Hard Ceramic Composites.

Diamond-silicon carbide (SiC) polycrystalline composite blends are studied using a computational approach combining molecular dynamics (MD) simulations for obtaining grain boundary (GB) fracture properties and phase field mechanics for capturing polycrystalline deformation and failure. An authentic microstructure, reconstructed from experimental lattice diffraction data with locally refined discretization in GB regions, is used to probe effects of local heterogeneities on material response in phase field simulations. The nominal microstructure consists of larger diamond and SiC (cubic polytype) grains, a matrix of smaller diamond grains and nanocrystalline SiC, and GB layers encasing the larger grains. These layers may consist of nanocrystalline SiC, diamond, or graphite, where volume fractions of each phase are varied within physically reasonable limits in parametric studies. Distributions of fracture energies from MD tension simulations are used in the phase field energy functional for SiC-SiC and SiC-diamond interfaces, where grain boundary geometries are obtained from statistical analysis of lattice orientation data on the real microstructure. An elastic homogenization method is used to account for distributions of second-phase graphitic inclusions as well as initial voids too small to be resolved individually in the continuum field discretization. In phase field simulations, SiC single crystals may twin, and all phases may fracture. The results of MD calculations show mean strengths of diamond-SiC interfaces are much lower than those of SiC-SiC GBs. In phase field simulations, effects on peak aggregate stress and ductility from different GB fracture energy realizations with the same mean fracture energy and from different random microstructure orientations are modest. Results of phase field simulations show unconfined compressive strength is compromised by diamond-SiC GBs, graphitic layers, graphitic inclusions, and initial porosity. Explored ranges of porosity and graphite fraction are informed by physical observations and constrained by accuracy limits of elastic homogenization. Modest reductions in strength and energy absorption are witnessed for microstructures with 4% porosity or 4% graphite distributed uniformly among intergranular matrix regions. Further reductions are much more severe when porosity is increased to 8% relative to when graphite is increased to 8%.

Modeling lung tissue dynamics and injury under pressure and impact loading.

A nonlinear viscoelastic model for the lung is implemented and evaluated for high-rate loading. Principal features of the model include a closed-cell approximation of the bulk compressibility accounting for air inside the lung and a damage-injury component by which local trauma is induced by cumulative normalized internal energy and amplified by gradients of energy density. The latter feature is adapted for use in standard numerical (i.e., explicit finite element) simulations in terms of the local rate of strain energy density and the longitudinal wave speed. Injury predictions for direct loading of a block of extracted lung material, rather than the entire thorax, via pressure pulses are in reasonably close agreement with experimental observations for an extracted rabbit lung: a threshold applied pressure exists above which edema is observed experimentally, correlating with low but non-negligible damage in the numerical results. Responses to impact by cylindrical and spherical projectiles are also interrogated. Penetration depths are comparable to those observed experimentally, as is drastically increasing damage with increasing impact velocity. Damage initiates and propagates from the impact surface, with local severity of injury decreasing with distance from the impact zone, in agreement with some empirical evidence. The model predicts more severe local injury, relative to the aforementioned surface pressure loading, than what is observed experimentally. Possible reasons for the discrepancy are analyzed, and adjustments to the model, with caveats, are suggested accordingly.

Expression and function of the Drosophila ACT88F actin isoform is not restricted to the indirect flight muscles.

Most higher eukaryotic genomes contain multiple actin genes, yet the sequence differences between isoforms are few. In Drosophila melanogaster it was previously established that one of the six actin genes, Act88F, is expressed only in the indirect flight muscles (IFMs). These muscles are highly specialised for oscillatory contractions to power flight. The implication was that this isoform had tissue-specific properties. In this paper we show using two reporter constructs expressing either beta-galactosidase, Act88F-lacZ, or the green fluorescent protein, Act88F-GFP, that the Act88F promoter is active in a small number of other muscles, including leg (femoral) and uterine muscles. However, the levels of Act88F driven non-IFM expression are much less than in the IFMs. We have confirmed endogenous Act88F gene expression in these other muscles by in situ hybridisation studies. Using null and antimorphic mutants to show decreased walking ability and delayed/reduced oviposition we demonstrated that Act88F expression is functionally important in multiple muscle groups. Since the mutant effects are mild, this supports the expectation that other actin genes are also expressed in these muscles. The Act88F-GFP promoter-reporter also detects Act88F-driven expression in the bristle-forming cells in the pupal wings. The implications of these results for the functions and developmental expression of the Drosophila ACT88F isoform are discussed.

Crystallization and preliminary X-ray analysis of Drosophila glutathione S-transferase-2.

The sigma-class glutathione S-transferase-2 (GST-2) from Drosophila melanogaster is predominantly found within the indirect flight muscles (IFMs), where it is bound to the 'heavy' subunit of the IFM thin filament troponin complex (Tn-H). An N-terminal extension found in GST-2 is unique within the sigma GST class and may be involved in its interaction with Tn-H or modulate its enzymatic function. The recombinant protein has been crystallized at room temperature using ammonium sulfate as precipitant. Synchrotron radiation was used to measure a complete native data set to 1.75 A resolution from flash-cooled crystals. The crystals belong to one of the trigonal space groups P3(1)21 or P3(2)21, with unit-cell parameters a = b = 89.7, c = 131.8 A. The self-rotation function is consistent with a GST-2 dimer in the asymmetric unit.

Keeping time with the human genome.

The cloning and characterization of 'clock gene' families has advanced our understanding of the molecular control of the mammalian circadian clock. We have analysed the human genome for additional relatives, and identified new candidate genes that may expand our knowledge of the molecular workings of the circadian clock. This knowledge could lead to the development of therapies for treating jet lag and sleep disorders, and add to our understanding of the genetic contribution of clock gene alterations to sleep and neuropsychiatric disorders. The human genome will also aid in the identification of output genes that ultimately control circadian behaviours.

The clock gene period of the housefly, Musca domestica, rescues behavioral rhythmicity in Drosophila melanogaster. Evidence for intermolecular coevolution?

In Drosophila, the clock gene period (per), is an integral component of the circadian clock and acts via a negative autoregulatory feedback loop. Comparative analyses of per genes in insects and mammals have revealed that they may function in similar ways. However in the giant silkmoth, Antheraea pernyi, per expression and that of the partner gene, tim, is not consistent with the negative feedback role. As an initial step in developing an alternative dipteran model to Drosophila, we have identified the per orthologue in the housefly, Musca domestica. The Musca per sequence highlights a pattern of conservation and divergence similar to other insect per genes. The PAS dimerization domain shows an unexpected phylogenetic relationship in comparison with the corresponding region of other Drosophila species, and this appears to correlate with a functional assay of the Musca per transgene in Drosophila melanogaster per-mutant hosts. A simple hypothesis based on the coevolution of the PERIOD and TIMELESS proteins with respect to the PER PAS domain can explain the behavioral data gathered from transformants.

Interaction of troponin-H and glutathione S-transferase-2 in the indirect flight muscles of Drosophila melanogaster.

Drosophila indirect flight muscles (IFMs) contain a 35 kDa protein which cross-reacts with antibodies to the IFM specific protein troponin-H isoform 34 (TnH-34). Peptide fingerprinting and peptide sequencing showed that this 35 kDa protein is glutathione S-transferase-2 (GST-2). GST-2 is present in the asynchronous indirect flight muscles but not in the synchronous tergal depressor of the trochanter (jump muscle). Genetic dissection of the sarcomere showed that GST-2 is stably associated with the thin filaments but the presence of myosin is required to achieve the correct stoichiometry, suggesting that there is also an interaction with the thick filament. The two Drosophila TnHs (isoforms 33 and 34) are naturally occurring fusion proteins in which a proline-rich extension of approximately 250 amino acids replaces the 27 C-terminal residues of the muscle-specific tropomyosin II isoform. The proteolytic enzyme, Igase, cleaves the hydrophobic C-terminal sequence of TnH-34 at three sites and TnH-33 at one site. This results in the release of GST-2 from the myofibril. The amount of GST-2 stably bound to the myofibril is directly proportional to the total amount of undigested TnH. It is concluded that GST-2 in the thin filament is stabilized there by interaction with TnH. We speculate that the hydrophobic N-terminal region of GST-2 interacts with the hydrophobic C-terminal extension of TnH, and that both are close to a myosin cross-bridge.

Drosophila actin mutants and the study of myofibrillar assembly and function.

The use of Drosophila mutations in the indirect flight muscle-specific actin gene, Act88F, to study actin structure/function and its assembly into thin filaments during myofibrillogenesis is described. Mutants with different phenotypic effects are discussed and attempts made to correlate the different properties of the mutants in vivo-myofibrillar structure, actin synthesis, accumulation and stability, heat shock response induction-with properties of the same mutations expressed by in vitro transcription/translation of the cloned actin genes-co-polymerisation, thermostability and protein conformation. Few of the properties show a complete correlation between the different classes of mutants. The nature of the diversity of the mutant effects is discussed. Questions as to how this will help in elucidating the molecular effects of the mutations and the assembly of thin filaments and myofibrils are considered. In addition, the efficacy of the co-polymerisation assay is examined. The post-translational processing of this actin-by N-terminal processing, methylation and ubiquitination-are described. Data is presented that inhibition of the N-terminal processing of actin in vitro affects the ability of the actin to copolymerise, and makes unprocessed actin behave as a capping protein. The possible in vivo importance of this phenomenon is discussed.

Inhibition of utilization of hypoxanthine and guanine in cells treated with the carbocyclic analog of adenosine. Phosphates of carbocyclic nucleoside analogs as inhibitors of hypoxanthine (guanine) phosphoribosyltransferase.

In cell cultures treated with the carbocyclic analog of adenosine (C-Ado, (+/-)-aristeromycin), the utilization of hypoxanthine and guanine has been observed to be blocked. In an attempt to define the mechanism of this inhibition, we have reexamined the metabolism of C-Ado and its effects on the metabolism of guanine and hypoxanthine. In cultures of L1210 cells, C-Ado at a concentration of 25 microM inhibited the utilization of hypoxanthine and guanine for nucleotide synthesis by more than 90% but produced little or no inhibition of the utilization of these bases in cultures of L1210/MeMPR cells which lack adenosine kinase and cannot phosphorylate C-Ado. In cultures of mammalian cells (L1210, HEp-2, and colon-26 cells), C-Ado was converted to the triphosphate (as previously observed) and also to the triphosphate of the carbocyclic analog of guanosine. The presence of coformycin in the medium at a concentration sufficient to inhibit AMP deaminase almost completely prevented the formation of carbocyclic GTP; thus, the deamination of C-Ado monophosphate is essential for the formation of phosphates of carbocyclic guanosine. Since hypoxanthine (guanine) phosphoribosyltransferase is known to be subject to end product inhibition, it was considered likely that phosphates of carbocyclic guanosine or carbocyclic inosine, present in C-Ado-treated cells, were responsible for inhibition of utilization of hypoxanthine and guanine. The 5'-phosphates of the carbocyclic analogs of inosine and guanosine were synthesized and found to be effective inhibitors of the phosphoribosyltransferase. Carbocyclic GMP was a better inhibitor than carbocyclic IMP and was also superior to GMP and IMP; the concentration of C-GMP that produced a 50% inhibition of GMP formation was approximately 1 microM. It is probable that the presence of phosphates of carbocyclic guanosine accounts for the inhibition of utilization of hypoxanthine and guanine in C-Ado-treated cells.

Synthesis and antiviral evaluation of carbocyclic analogues of ribofuranosides of 2-amino-6-substituted-purines and of 2-amino-6-substituted-8-azapurines.

Carbocyclic analogues of ribofuranosides of 2-amino-6-substituted-purines and of 2-amino-6-substituted-8- azapurines were prepared from the 2-amino-6-chloropurine ribofuranoside analogue (2) and the 2-amino-6-chloro-8- azapurine ribofuranoside analogue (9), respectively. Analogues of purine ribofuranosides with the chloro, amino, methylamino, or methylthio group at position 6, the thioguanosine analogue, and the previously reported guanosine analogue were evaluated in vitro against herpes simplex virus, type 1 (HSV-1). 8- Azapurine ribofuranoside analogues with the chloro, amino, or methylthio group at position 6 and the previously reported 8- azaguanosine analogue were also evaluated against HSV-1. The carbocyclic analogue (6) of 2,6-diaminopurine ribofuranoside is highly active against HSV-1 and, also, against vaccinia virus. The 2-amino-6-chloropurine, 2-amino-6-(methylamino)purine, and the 2,6-diamino-8- azapurine derivatives also demonstrated significant activity against HSV-1.