Realization of couplings in a polynomial type mixed-mode cellular neural network

In this paper realization of couplings between cells in a polynomial type mixed-mode cellular neural network (CNN) is analyzed. The choice of the multiplier is discussed and two multiplier types are analyzed. Also, two circuits for generating the second and third order polynomial terms of cell output are described. The accuracy of the multipliers and polynomial circuits at the presence of device mismatch is analyzed.     

Filamentous ascomycetes inhabiting the rhizoid environment of the liverwort Cephaloziella varians in Antarctica are assessed by direct PCR and cloning

We molecularly assesed the ascomycetous fungal communities inhabiting the rhizoid environment of Cephaloziella varians, collected at Rothera Point on the western Antarctic Peninsula. The RFLP-phenotyped and cloned PCR products of a partial small subunit of the ribosomal RNA gene were sequenced and analyzed with neighbor joining (NJ) and maximum parsimony (MP). Both analyses identified four bootstrap-supported groups: (i) a sister group to Onygenales, (ii) a well-supported clade with Phialocephala fortinii, (iii) a large group of clones nested within Chaetothyriales, and (iv) a group nested within Eurotiales with a likely affinity to the genus Aspergillus. An additional marginally supported clade, including helotialean Hymenoscyphus fructigenus, was detected in the NJ analysis. Placement of one clone (possibly helotialean) was not supported by either analysis. We included Hymenoscyphus ericae (Helotiales) in our analyses to test for its presence in clone libraries. None of our clones showed strongly supported affinity to H. ericae. The culture-independent technique proved useful for assessing the composition of rhizoid fungal communities, although it remains unknown whether any of these fungi colonize C. varians tissues. Direct-community assays of this kind might be best combined with traditional isolation techniques to get a more holistic view of fungi occupying plant tissues.     

Discovery in silico and characterization in vitro of novel genes exclusively expressed in the mouse epididymis

Epididymal proteins interact with sperm during their passage through the epididymis and thus contribute to the maturation and fertilizing capacity of the spermatozoa. In the present study we have discovered five novel epididymis-specific genes through in silico analysis of expressed sequence tags (ESTs) at the UniGene library collection. The strategy used is a powerful way to discover novel epididymis-specific genes. The full-length cDNA sequences were determined, and computational tools were used to characterize the genomic structures and to predict putative functions for the encoded proteins. In vitro analyses revealed that all five genes characterized were highly expressed in the defined areas of the epididymis, and they were not expressed at significant levels in any other tissue. Three of the genes were named on the basis of their putative functions: Spint4 (serine protease inhibitor, Kunitz type 4), and Rnase9 and Rnase10 (ribonuclease, Rnase A family 9 and 10), while for the ESTs AV381130 and AV381126 no putative functions could be predicted. The expression of Spint4, Rnase9, and AV381130 was found to be under a direct or indirect regulation by androgens, while the expression of Rnase10 is regulated by a testicular factor(s) other than androgen. None of the genes were expressed in the immature epididymis, while mRNAs were detected from d 17 onward, at the time of maturation of epididymal epithelium. However, the expression of AV381130 was not detected until d 30 after birth, indicating a close connection between gene expression and puberty.     

Insider information: what viruses tell us about endocytosis

Viruses have long served as tools in molecular and cellular biology to study a variety of complex cellular processes. Currently, there is a revived interest in virus entry into animal cells because it is evident that incoming viruses make use of numerous endocytic pathways that are otherwise difficult to study. Besides the classical clathrin-mediated uptake route, viruses use caveolae-mediated endocytosis, lipid-raft-mediated endocytic pathways, and macropinocytosis. Some of these are subject to regulation, involve novel endocytic organelles, and some of them connect organelles that were previously not known to communicate by membrane traffic.     

Monastrol, a prototype anti-cancer drug that inhibits a mitotic kinesin, induces rapid bursts of axonal outgrowth from cultured postmitotic neurons

Terminally postmitotic neurons continue to express many of the kinesin-related proteins known to configure microtubules during mitosis. Drugs that inhibit these kinesins are being developed as anti-cancer agents with the hope that they will inhibit proliferation of tumor cells without having adverse effects on the nervous system. The prototype, termed monastrol, inhibits the kinesin known as Eg5, which is essential for maintaining separation of the half-spindles. Eg5 is also highly expressed in neurons, particularly during development. Exposure of cultured sympathetic neurons to monastrol for a few hours increased both the number and the growth rate of the axons. With additional time, the overall lengths of the axons were indistinguishable from controls. Sensory neurons showed a similar short-term increase in axonal growth rate. However, prolonged exposure resulted in shorter axons, suggesting that sensory neurons may be more sensitive to toxic effects of the drug. Nevertheless, the overall health of the cultures was still far more robust than cultures treated with taxol, a drug commonly used for anti-cancer therapy. On the basis of these results, we conclude that Eg5 normally generates forces that oppose axonal growth, presumably by partially suppressing the forward advance of microtubules. We speculate that local regulation of Eg5 could be a means by which neurons coordinate rapid bursts of axonal growth with appropriate environmental cues. The comparatively modest toxic effects on the neurons over time are a hopeful sign for clinicians interested in using anti-Eg5 drugs for cancer therapy.     

Genetic polymorphisms of DNA repair and xenobiotic-metabolizing enzymes: effects on levels of sister chromatid exchanges and chromosomal aberrations

Elevated levels of chromosomal aberrations (CAs) in peripheral blood lymphocytes, widely used as a cytogenetic biomarker of genotoxic effects, have been linked to cancer predisposition. However, tobacco smoking, occupational carcinogen exposure, or time since CA analysis do not appear to explain the cancer predictivity of CAs. Alternatively, the observed CA-cancer association could reflect unidentified exposures or individual susceptibility. We assessed the effects of genetic polymorphisms of DNA repair proteins and xenobiotic-metabolizing enzymes (XMEs) on the levels of CAs and sister chromatid exchanges (SCEs) in peripheral lymphocytes of 145 (CAs) and 60 (SCEs) healthy Caucasians. Genotypes of DNA repair genes X-ray repair cross-complementation group 1 (XRCC1 codons 194, 280, 399) and 3 (XRCC3 codon 241 [corrected]), and XME genes glutathione-S-transferase (GST) M1 and T1 and N-acetyl transferase 2 (NAT2) were determined using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP)-based methods. After Poisson regression adjustment for age, sex, smoking, country, and genotypes, a higher frequency of chromosome-type breaks was observed for NAT2 slow acetylators (in nonsmokers) and GSTT1 null subjects (in smokers). Individuals carrying variant alleles for XRCC1 codons 280 and 194 showed a decreased level of chromosome-type breaks. The effect of GSTM1 null and XRCC1 codon 399 genotypes on the frequency of CAs was modified by smoking. In linear regression models adjusting for age, sex, smoking, and genotypes, none of the polymorphisms significantly affected SCE frequency, although GSTT1 null subjects had a slightly elevated SCE level. Our results are in line with earlier findings on the influence of NAT2, GSTT1, and GSTM1 polymorphisms on the level of lymphocyte chromosome damage and suggest that also XRCC1 polymorphism affects CA frequencies, thus apparently influencing DNA repair phenotype. It remains to be examined whether these or other genetic polymorphisms could explain the observed cancer risk predictivity of high CA frequency.     

Genomic overview of serine proteases

Serine proteases (SP) are peptidases with a uniquely activated serine residue in the substrate-binding pocket. They represent about 0.6% of all proteins in the human genome. SP are involved in many vital functions such as digestion, blood clotting, fibrinolysis, fertilization, and complement activation and are related to many diseases including cancer, arthritis, and emphysema. In this study, we performed a genomic analysis of human serine proteases utilizing different databases, primarily that of MEROPS. SP are distributed along all human chromosomes except 18 and Y with the highest density (23 genes) on chromosome 19. They are either randomly located within the genome or occur in clusters. We identified a number of SP clusters, the largest being the kallikrein cluster on chromosome 19q13.4 which is formed of 15 adjacent genes. Other clusters are located on chromosomes 19p13, 16p13, 14q11, 13q35, 11q22, and 7q35. Genes of each cluster tend to be of comparable sizes and to be transcribed in the same direction. The members of some clusters are sometimes functionally related, e.g., the involvement of many kallikreins in endocrine-related malignancies and the hematopoietic cluster on chromosome 14. It is hypothesized that members of some clusters are under common regulatory mechanisms and might be involved in cascade enzymatic pathways. Several functional domains are found in SP, which reflect their functional diversity. Membrane-type SP tend to cluster in 3 chromosomes and have some common structural domains. Several databases are available for screening, structural and functional analysis of serine proteases. With the near completion of the Human Genome Project, research will be more focused on the interactions between SP and their involvement in pathophysiological processes.     

Quality control in the endoplasmic reticulum

The endoplasmic reticulum (ER) has a quality-control system for 'proof-reading' newly synthesized proteins, so that only native conformers reach their final destinations. Non-native conformers and incompletely assembled oligomers are retained, and, if misfolded persistently, they are degraded. As a large fraction of ER-synthesized proteins fail to fold and mature properly, ER quality control is important for the fidelity of cellular functions. Here, we discuss recent progress in understanding the conformation-specific sorting of proteins at the level of ER retention and export.     

Regulation of ENA1 Na(+)-ATPase gene expression by the Ppz1 protein phosphatase is mediated by the calcineurin pathway

Saccharomyces cerevisiae strains lacking the Ppz1 protein phosphatase are salt tolerant and display increased expression of the ENA1 Na⁺-ATPase gene, a major determinant for sodium extrusion, while cells devoid of the similar Ppz2 protein do not show these phenotypes. However, a ppz1 ppz2 mutant displays higher levels of ENA1 expression than the ppz1 strain. We show here that the increased activity of the ENA1 promoter in a ppz1 ppz2 mutant maps to two regions: one region located at −751 to −667, containing a calcineurin-dependent response element (CDRE), and one downstream region (−573 to −490) whose activity responds to intracellular alkalinization. In contrast, the increased ENA1 expression in a ppz1 mutant is mediated solely by an intact calcineurin/Crz1 signaling pathway, on the basis that (i) this effect maps to a single region that contains the CDRE and (ii) it is blocked by the calcineurin inhibitor FK506, as well as by deletion of the CNB1 or CRZ1 gene. The calcineurin dependence of the increased ENA1 expression of a ppz1 mutant would suggest that Ppz1 could negatively regulate calcineurin activity. In agreement with this notion, a ppz1 strain is calcium sensitive, and this mutation does not result in a decrease in the calcium hypertolerance of a cnb1 mutant. It has been shown that ENA1 can be induced by alkalinization of the medium and that a ppz1 ppz2 strain has a higher intracellular pH. However, we present several lines of evidence that show that the gene expression profile of a ppz1 mutant does not involve an alkalinization effect. In conclusion, we have identified a novel role for calcineurin, but not alkalinization, in the control of ENA1 expression in ppz1 mutants.