CCL3 and viral chemokine-binding protein gg modulate pulmonary inflammation and virus replication during equine herpesvirus 1 infection

CCL3 is a proinflammatory chemokine that mediates many of the cellular changes occurring in pulmonary disease. Here, CCL3^−/− mice were used to investigate the role of this chemokine during respiratory herpesvirus infection. Compared to wild-type mice, CCL3^−/− mice infected with the alphaherpesvirus equine herpesvirus 1 (EHV-1) displayed reduced body weight loss but had higher pulmonary viral loads. Lungs from infected CCL3^−/− mice suffered a milder interstitial pneumonia, and fewer immune cells were recovered from the pulmonary airways after infection. We could also demonstrate that herpesvirus-encoded chemokine-binding glycoprotein G (gG) was capable of inhibiting the chemotactic functions of CCL3. This CCL3-mediated chemotaxis, however, was restored in the presence of gG-specific antibodies, which puts into question the advertised use of gG deletion mutants as marker vaccines. In summary, we concluded that CCL3 is a major player in controlling herpesvirus replication in the target organ, the lung, and does so by evoking a strong inflammatory response. The immunomodulatory activity of CCL3 is balanced by the expression of viral gG, whose chemokine-binding activity is mitigated in secondary infections by the production of anti-gG antibodies.     

Immunohistochemical Localization of Manserin,a Novel Neuropeptide Derived from Secretogranin II,in Rat Adrenal Gland,and its Upregulation by Physical Stress

We recently identified a novel 40-amino acid neuropeptide designated manserin from the rat brain (Yajima in NeuroReport 15: 1755–1759, 2004). Manserin is highly expressed in pituitary and hypothalamic nuclei, which suggests that it plays a role in the endocrine system. In this study, we employed immunohistochemical methods to investigate the presence of manserin in rat adrenal glands, as well as its regulation by physical stress. Immunohistochemical analysis using anti-manserin antibody showed that manserin is present in the rat adrenal medulla but not in the cortex. When the colocalization of manserin and phenylethanolamine N-methyltransferase (PNMT), an epinephrine-synthesizing enzyme, was examined, virtually all PNMT-positive cells expressed manserin. Interestingly, the immunoreactivity of manserin was significantly increased when the rats were exposed to water-immersion restraint stress. These results demonstrate for the first time that adrenal manserin, a novel neuropeptide, may have a potential physiological role under stress-inducing conditions.     

Systematic separation and purification of elastase, gelatinase (matrix metalloproteinase 9), and collagenase (matrix metalloproteinase 8) from polymorphonuclear leukocytes in dialyzers previously used by patients with renal failure

We developed a simple and effective method for the systematic separation and purification of human polymorphonuclear leukocyte (PMN) proteinases, elastase, gelatinase (matrix metalloproteinase 9, type IV collagenase), and collagenase (matrix metalloproteinase 8), derived from the extracts of hollow fiber dialyzers that had been utilized in the treatment of patients with renal failure. The fraction containing elastase was grossly separated from that containing gelatinase and collagenase by heparin-Sepharose chromatography and purified in an aprotinin column. The remaining two enzymes were then separated using the gelatin-Sepharose column after gel chromatography following ammonium sulfate precipitation. Gelatinase and collagenase were further purified by gelatin-Sepharose chromatography as a latent form and by collagen-Sepharose chromatography as an activated form. This novel method offers procedural advantages over existing methods that separate PMNs from the whole blood of volunteers for experimental research purposes.     

Ornithine decarboxylase antizyme upregulates DNA-dependent protein kinase and enhances the nonhomologous end-joining repair of DNA double-strand breaks in human oral cancer cells

Ornithine decarboxylase (ODC) antizyme targets ODC for ubiquitin-independent proteosome degradation, thereby inhibiting polyamine synthesis. It has been shown to regulate DNA methylation and has tumor suppressor activity. Increasing evidence suggested that antizyme may also have ODC-independent functions. Here, we report that antizyme plays a role in DNA double-strand break repairs. A zinc-inducible human antizyme gene expression vector was transfected into UM1 human oral squamous cancer cells that do not express endogenous antizyme. The resultant upregulated genes were screened by cDNA arrays and confirmed by quantitative real-time polymerase chain reaction. DNA-dependent protein kinase including its catalytic subunit DNA-PKcs and regulatory subunit Ku70, two key proteins of the DNA damage repair machinery, was significantly upregulated after ectopic expression of antizyme. Consistently, we found that UM1 cells are sensitive to gamma irradiation and deficient in DNA damage repairs, as shown by radio-sensitivity and Comet assays. Ectopic expression of antizyme increased radio-resistance of UM1 cells and restored their capacity of DNA damage repairs to the level of UM2 cells that have an identical genetic background but express endogenous antizyme. Plasmid end-joining assays confirmed that antizyme enhances the ability of UM1 cells to repair DNA double-strand breaks by the nonhomologous end-joining pathway.     

Inference of S-system models of genetic networks using a cooperative coevolutionary algorithm

MOTIVATION: To resolve the high-dimensionality of the genetic network inference problem in the S-system model, a problem decomposition strategy has been proposed. While this strategy certainly shows promise, it cannot provide a model readily applicable to the computational simulation of the genetic network when the given time-series data contain measurement noise. This is a significant limitation of the problem decomposition, given that our analysis and understanding of the genetic network depend on the computational simulation. RESULTS: We propose a new method for inferring S-system models of large-scale genetic networks. The proposed method is based on the problem decomposition strategy and a cooperative coevolutionary algorithm. As the subproblems divided by the problem decomposition strategy are solved simultaneously using the cooperative coevolutionary algorithm, the proposed method can be used to infer any S-system model ready for computational simulation. To verify the effectiveness of the proposed method, we apply it to two artificial genetic network inference problems. Finally, the proposed method is used to analyze the actual DNA microarray data.     

Two forms of secreted and thermostable luciferases from the marine copepod crustacean, Metridia pacifica

We cloned two forms of the secreted and thermostable luciferase genes, MpLuc1 and MpLuc2, from the marine copepod, Metridia pacifica. The 840-bp MpLuc1 cDNA comprised a 630-bp open reading frame encoding a 210-amino acid polypeptide (22.7 kDa). MpLuc1 had the closest homology with Metridia longa luciferase. The 753-bp MpLuc2 cDNA consisted of a 567-bp open reading frame (20.3 kDa), and it had the closest homology with Gaussia princeps luciferase. Single-specimen genomic PCR confirmed the presence of two luciferase genes in M. pacifica, and single-specimen RT-PCR revealed that both luciferase mRNAs were expressed. Both MpLuc1 and MpLuc2 (MpLucs) specifically reacted with the substrate coelenterazine producing identical bioluminescent spectra (lambdamax, 485 nm), but with different kinetics. Adding salt such as MgCl2 and CaCl2 to the reaction mixture significantly enhanced MpLuc1 and MpLuc2 activities. Wild-type MpLucs were remarkably thermostable; MpLuc1 retained about 60% of the original activity even after incubation at 90 degrees C for 30 min. MpLucs expressed in NIH-3T3 and HeLa cells were largely secreted into the culture medium. Continuous monitoring of secreted MpLuc1 driven by the c-fos promoter demonstrated the potential usefulness of MpLuc1 in nondisruptive reporter assays.     

Evolution of Pulmonate Gastropod Mitochondrial Genomes: Comparisons of Gene Organizations of Euhadra, Cepaea and Albinaria and Implications of Unusual Trna Secondary Structures

Complete gene organizations of the mitochondrial genomes of three pulmonate gastropods, Euhadra herklotsi, Cepaea nemoralis and Albinaria coerulea, permit comparisons of their gene organizations. Euhadra and Cepaea are classified in the same superfamily, Helicoidea, yet they show several differences in the order of tRNA and protein coding genes. Albinaria is distantly related to the other two genera but shares the same gene order in one part of its mitochondrial genome with Euhadra and in another part with Cepaea. Despite their small size (14.1-14.5 kbp), these snail mtDNAs encode 13 protein genes, two rRNA genes and at least 22 tRNA genes. These genomes exhibit several unusual or unique features compared to other published metazoan mitochondrial genomes, including those of other molluscs. Several tRNAs predicted from the DNA sequences possess bizarre structures lacking either the T stem or the D stem, similar to the situation seen in nematode mt-tRNAs. The acceptor stems of many tRNAs show a considerable number of mismatched basepairs, indicating that the RNA editing process recently demonstrated in Euhadra is widespread in the pulmonate gastropods. Strong selection acting on mitochondrial genomes of these animals would have resulted in frequent occurrence of the mismatched basepairs in regions of overlapping genes.     

Neuromechanics: an integrative approach for understanding motor control

Neuromechanics seeks to understand how muscles, sense organs,motor pattern generators, and brain interact to produce coordinatedmovement, not only in complex terrain but also when confrontedwith unexpected perturbations. Applications of neuromechanicsinclude ameliorating human health problems (including prosthesisdesign and restoration of movement following brain or spinalcord injury), as well as the design, actuation and control ofmobile robots. In animals, coordinated movement emerges fromthe interplay among descending output from the central nervoussystem, sensory input from body and environment, muscle dynamics,and the emergent dynamics of the whole animal. The inevitablecoupling between neural information processing and the emergentmechanical behavior of animals is a central theme of neuromechanics.Fundamentally, motor control involves a series of transformationsof information, from brain and spinal cord to muscles to body,and back to brain. The control problem revolves around the specifictransfer functions that describe each transformation. The transferfunctions depend on the rules of organization and operationthat determine the dynamic behavior of each subsystem (i.e.,central processing, force generation, emergent dynamics, andsensory processing). In this review, we (1) consider the contributionsof muscles, (2) sensory processing, and (3) central networksto motor control, (4) provide examples to illustrate the interplayamong brain, muscles, sense organs and the environment in thecontrol of movement, and (5) describe advances in both roboticsand neuromechanics that have emerged from application of biologicalprinciples in robotic design. Taken together, these studiesdemonstrate that (1) intrinsic properties of muscle contributeto dynamic stability and control of movement, particularly immediatelyafter perturbations; (2) proprioceptive feedback reinforcesthese intrinsic self-stabilizing properties of muscle; (3) controlsystems must contend with inevitable time delays that can simplifyor complicate control; and (4) like most animals under a varietyof circumstances, some robots use a trial and error processto tune central feedforward control to emergent body dynamics.