DNA sequence evidence for polymorphic forms of human serum amyloid A (SAA)

Serum amyloid A (SAA) is an acute-phase reactant and precursor to amyloid A protein, the major constituent of the fibril deposits of reactive amyloidosis. The factors determining whether the 104-amino acid SAA molecule is converted into the 76-amino acid amyloid A protein and deposited as fibrils are not known. As an initial step toward investigating the possibility that a particular primary structure of SAA is involved in amyloid formation, we have cloned and determined the nucleotide sequence of human SAA-specific cDNAs. The first clone, selected using an oligonucleotide probe, was shown to encode the signal peptide and amino-terminal region of SAA. The cDNA of this clone served as probe in the selection of two distinct, full-length SAA cDNAs, initially differentiated by the presence (pSAA21) or absence (pSAA82) of a PstI site in the coding sequence. The complete nucleotide sequence of pSAA82 cDNA was determined. Since there appear to be multiple human SAA alleles, it is conceivable that their differential expression is important to amyloid formation.     

Monoclonal antibody that inhibits infection of HeLa and rhabdomyosarcoma cells by selected enteroviruses through receptor blockade.

BALB/c mice were immunized with HeLa cells, and their spleen cells were fused with myeloma cells to produce hybridomas. Initial screening of culture fluids from 800 fusion products in a cell protection assay against coxsackievirus B3 (CB3) and the CB3-RD virus variant yielded five presumptive monoclonal antibodies with three specificities: protection against CB3 on HeLa, protection against CB3-RD on rhabdomyosarcoma (RD) cells, and protection against both viruses on the respective cells. Only one of the monoclonal antibodies (with dual specificity) survived two subclonings and was studied in detail. The antibody was determined to have an immunoglobulin G2a isotype and protected cells by blockade of cellular receptors, since attachment of [35S]methionine-labeled CB3 was inhibited by greater than 90%. The monoclonal antibody protected HeLa cells against infection by CB1, CB3, CB5, echovirus 6, and coxsackievirus A21 and RD cells against CB1-RD, CB3-RD, and CB5-RD virus variants. The monoclonal antibody did not protect either cell type against 16 other immunotypes of picornaviruses. The monoclonal antibody produced only positive fluorescence on those cells which were protected against infection, and 125I-labeled antibody confirmed the specific binding to HeLa and RD cells. The results suggest that this monoclonal antibody possesses some of the receptor specificity of the group B coxsackieviruses.     

Allometry in the placoderm Bothriolepis canadensis and its significance to antiarch evolution

A set of 18 measurements of the dermal armour of Bothriolepis canadensis Whiteaves (Placodermi, Anti-archa) is analysed with respect to allometric growth patterns. The strongest allometric patterns were found for the orbital fenestra and premedian plate of the head-shield. and the anterior median dorsal plate of the trunk-shield. These are all areas of the greatest importance in antiarch phylogeny and imply a role for ontogenetic effects such as paedomorphosis in the evolution of antiarchs. It is suggested that this is partly a result of the severe constraints on growth in a closed box such as the armour of placoderms, and may be generally true of such arrangements.     

Sapwood area ofPinus contorta stands as a function of mean size and density

Summary An indirect test of the relationship between leaf area and the combination of mean size and density is made in stands of lodgepole pine (Pinus contorta Dougl.). Total sapwood cross-sectional area of these stands is a function of the product of density and mean diameter raised to an exponent of about 1.6. Results from other studies, representing four species, suggest that this relationship between sapwood area and the combination of mean size and density may be general. The implications of the relationship are discussed in the context of evapotranspiration, competition and self-thinning.Utah Agricultural Experiment Station Journal Paper No. 3072     

Induction of endogenous xenotropic retrovirus expression by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate

Summary The tumor promotor 12-O-tetradecanoylphorbol-13-acetate (TPA) was examined for its ability to induce endogenous retrovirus from a high-passage clone of Kirsten sarcoma virus-transformed Balb/c (K-Balb) mouse cells. TPA activated virus in a concentration-dependent manner (0.0016 to 4.0 μM). Exposure to 1mM actinomycin D inhibited virus induction, suggesting that cellular RNA synthesis is required de novo by this inducer. A broad-spectrum neutralizing antibody to murine type C virus, gp70, was shown to neutralize the infectivity of the induced virus. The activated virus had the host range of the xenotropic Balb virus:2, and after removal of the inducer, the activated state decayed rapidly. TPA stimulated DNA, RNA, and protein synthesis in K-Balb cells, indicating that the mechanism of inducation may be different from that of previously identified virus inducers. The effects observed using the well-defined K-Balb system offer an opportunity to study the modulation of retrovirus gene expression by TPA. This work was conducted while the authors were with the Biological Carcinogenesis Program, Frederick Cancer Research Facility, Frederick, MD 21701, and was supported under Contract NO1-CO-75380 with the National Cancer Institute, National Institutes of Health, Bethesda, MD 20205.     

Conserved Nodulation Genes in Rhizobium meliloti and Rhizobium trifolii

Plasmids which contained wild-type or mutated Rhizobium meliloti nodulation (nod) genes were introduced into Nod⁻R. trifolii mutants ANU453 and ANU851 and tested for their ability to nodulate clover. Cloned wild-type and mutated R. meliloti nod gene segments restored ANU851 to Nod⁺, with the exception of nodD mutants. Similarly, wild-type and mutant R. meliloti nod genes complemented ANU453 to Nod⁺, except for nodCII mutants. Thus, ANU851 identifies the equivalent of the R. meliloti nodD genes, and ANU453 specifies the equivalent of the R. meliloti nodCII genes. In addition, cloned wild-type R. trifolii nod genes were introduced into seven R. meliloti Nod⁻ mutants. All seven mutants were restored to Nod⁺ on alfalfa. Our results indicate that these genes represent common nodulation functions and argue for an allelic relationship between nod genes in R. meliloti and R. trifolii.     

Rhizobium meliloti nodulation genes: identification of nodDABC gene products, purification of nodA protein, and expression of nodA in Rhizobium meliloti.

A set of conserved, or common, bacterial nodulation (nod) loci is required for host plant infection by Rhizobium meliloti and other Rhizobium species. Four such genes, nodDABC, have been indicated in R. meliloti 1021 by genetic analysis and DNA sequencing. An essential step toward understanding the function of these genes is to characterize their protein products. We used in vitro and maxicell Escherichia coli expression systems, together with gel electrophoresis and autoradiography, to detect proteins encoded by nodDABC. We facilitated expression of genes on these DNA fragments by inserting them downstream of the Salmonella typhimurium trp promoter, both in colE1 and incP plasmid-based vectors. Use of the incP trp promoter plasmid allowed overexpression of a nodABC gene fragment in R. meliloti. We found that nodA encodes a protein of 21 kilodaltons (kDa), and nodB encodes one of 28 kDa; the nodC product appears as two polypeptide bands at 44 and 45 kDa. Expression of the divergently read nodD yields a single polypeptide of 33 kDa. Whether these represent true Rhizobium gene products must be demonstrated by correlating these proteins with genetically defined Rhizobium loci. We purified the 21-kDa putative nodA protein product by gel electrophoresis, selective precipitation, and ion-exchange chromatography and generated antiserum to the purified gene product. This permitted the immunological demonstration that the 21-kDa protein is present in wild-type cells and in nodB- or nodC-defective strains, but is absent from nodA::Tn5 mutants, which confirms that the product expressed in E. coli is identical to that produced by R. meliloti nodA. Using antisera detection, we found that the level of nodA protein is increased by exposure of R. meliloti cells to plant exudate, indicating regulation of the bacterial nod genes by the plant host.     

Effects of histamine on bronchial artery blood flow and bronchomotor tone

The effects of aerosolized 5% histamine (10 breaths) on bronchial artery blood flow (Qbr), airflow resistance (RL), and pulmonary and systemic hemodynamics were studied in mechanically ventilated sheep anesthetized with pentobarbital sodium. Histamine increased mean Qbr and RL to 252 +/- 45 and 337 +/- 53% of base line, respectively. This effect was significantly different from base line for 30 min after challenge. The histamine-induced increase in RL was blocked by pretreatment with the histamine H1 receptor antagonist, chlorpheniramine, whereas the histamine-induced elevation in Qbr was prevented by the H2 antagonist, metiamide. Both responses were blocked only when both antagonists were present. Changes in Qbr were not directly associated with alterations in systemic and pulmonary hemodynamics or arterial blood gas composition. In vitro histamine caused a dose-dependent contraction of ovine bronchial artery strips that was prevented by H1 antagonist. The H2 agonist, impromidine, caused relaxation of precontracted arterial strips and was more potent and efficacious than histamine, whereas H1 agonists failed to elicit a relaxant response. Thus these findings indicate that histamine aerosol induces a vasodilation in the bronchial vascular bed; histamine has a direct effect on Qbr that is independent of alterations in RL, systemic and pulmonary hemodynamics, or arterial blood gas composition; and, histamine-induced bronchoconstriction is mediated predominantly by H1-receptors, whereas increased Qbr is controlled predominantly by H2-receptors, probably located in resistance vessels. This local effect of histamine on Qbr may have important implications in the pathophysiology of bronchial asthma and pulmonary edema.     

Generalized transduction in Rhizobium meliloti.

Generalized transduction of Rhizobium meliloti 1021 was carried out by bacteriophage N3. Genetic markers on the chromosome and the pSym megaplasmid were transduced, along with markers on several IncP plasmids. Cotransduction between transposon Tn5 insertions and integrated recombinant plasmid markers permitted correlation of cotransductional frequencies and known physical distances. Bacteriophage N3 was capable of infecting several commonly used strains of R. meliloti.