SuperSAGE analysis of the <Emphasis Type="Italic">Nicotiana attenuata</Emphasis> transcriptome after fatty acid-amino acid elicitation (FAC): identification of early mediators of insect responses

Background  

Plants trigger and tailor defense responses after perception of the oral secretions (OS) of attacking specialist lepidopteran larvae. Fatty acid-amino acid conjugates (FACs) in the OS of the Manduca sexta larvae are necessary and sufficient to elicit the herbivory-specific responses in Nicotiana attenuata, an annual wild tobacco species. How FACs are perceived and activate signal transduction mechanisms is unknown.     

Relationships between the human pepsinogen DNA and protein polymorphisms.

Pepsinogens (PGA) are the inactive precursors of pepsin, the major acid protease found in the stomach. The PGA gene family exhibits polymorphic variation in human populations that can either be demonstrated by electrophoretic analysis of the proteins or by analysis of the respective genes with cDNA probes. Here, we describe the interrelationships between the most common pepsinogen protein phenotypes and the corresponding pepsinogen haplotypes (A, B, and C) containing different combinations of the PGA3, PGA4, and PGA5 genes. We propose that this unusual genetic variation involving haplotypes that contain three, two, and one genes, respectively, is the result of molecular evolution by gene duplication.     

Differential expression of metastasis-associated cell surface glycoproteins and mRNA in a murine large cell lymphoma

A metastatic variant cell subline of the Abelson virus-transformed murine large lymphoma/lymphosarcoma RAW117 has been selected in vivo ten times for liver colonization. Highly metastatic subline RAW117-H10 forms greater than 200 times as many gross surface liver tumor nodules as the parental line RAW117-P. Analysis of cellular proteins and glycoproteins indicates reduced expression of murine Moloney leukemia virus-associated p15, p30, and gp70, and increased expression of a sialoglycoprotein, gp150, in the highly metastatic H10 cells. Northern analyses of oncogene expression suggested that mRNA of various oncogenes was expressed equally or not expressed in the RAW117 cells of differing metastatic potential. Differential gene expression was examined using a cDNA library of 17,600 clones established from poly A+ mRNA isolated from H10 cells. The cDNA library was screened by the colony hybridization technique using probes made from both RAW117-P and -H10 cells. Approximately 99.5% of these cDNA clones were expressed identically in P and H10 cells. Of the few differentially expressed cDNA clones (approx. 150/17,600), one-half of these were identified as Moloney leukemia virus sequences in a separate probing with a radiolabeled Moloney leukemia virus probe. The remainder of the differentially expressed mRNA detected by colony hybridization of the cDNA library were expressed at higher levels (approx. 1/6) or lower levels (approx. 1/3) in the highly metastatic H10 cells.     

Thermal denaturation of calf thymus DNA: existence of a GC-richer fraction

In 2.5 x 10(-4)M EDTA buffer, the derivative melting curve of calf thymus DNA shows a major band at 47 degrees with a shoulder at about 54 degrees . The fraction of melting area of this shoulder is about 13%. For reconstituted polylysine-calf thymus DNA complexes, in addition to the melting of free DNA regions at about 50 degrees (T(m)) there is another melting at about 106 degrees (T(m)) of polylysine-bound regions. The melting band of the complex at T(m) is not symmetrical. As more polylysine is bound to DNA the melting amplitude is diminished greatly on the major band at 47 degrees but only slightly on the shoulder at 54 degrees . The insensitivity of this shoulder appears to result from the existence of a 13% fraction of calf thymus DNA containing 55% GC. It is not favorably bound by polylysine. It remains in the supernatant after centrifugation and melts at about 54-56 degrees . This conclusion is further supported by two facts: the reconstitution method provides a condition for selective binding of polylysine to AT-rich DNA, and it yields a fully symmetric melting band at T(m) for complexes of polylysine with homogeneous bacterial DNA such as the one from M. luteus.     

Serological reactions associated with the clumping factor of Staphylococcus aureus

Rotter, Joan (University of Oklahoma Medical Center, Oklahoma City), and Florene C. Kelly. Serological reactions associated with the clumping factor of Staphylococcus aureus. J. Bacteriol. 91:588-594. 1966.-Evidence that the substance which causes staphylococci to clump in the presence of fibrinogen (clumping factor) is antigenically similar in strains which are serologically diverse according to agglutination reactions has been obtained from fibrinogen-cell clumping inhibition tests. Antisera for clumping factor (CF)-positive strains inhibited the clumping reaction of all strains tested. After adsorption with homologous cells or with cells of other CF-positive strains, the antisera no longer inhibited clumping. When antisera were adsorbed with trypsin-treated, CF-positive cells, or with cells of CF-negative mutants, the ability to inhibit the clumping reaction persisted. Antibody to CF activity was not associated with coagulase. Latex coated with extracts derived from the cells of five CF-positive and six CF-negative strains was, in each instance, agglutinated by sera from rabbits immunized with CF-positive cells. After adsorption with trypsinized, CF-positive cells, antisera still agglutinated latex which had been treated with the CF-positive extracts, but not with the CF-negative extracts. Similar results were obtained after antisera were adsorbed with the cells of CF-negative mutants. Cell agglutination titers of sera from rabbits immunized with CF-negative staphylococci were significantly lower than those produced in response to CF-positive cells, regardless of their coagulase activity. If the CF-inhibiting antibody also functions as an agglutinin, it apparently is not solely responsible for this difference.     

Accumulation of wild-type p53 protein upon gamma-irradiation induces a G2 arrest-dependent immunoglobulin kappa light chain gene expression.

The exposure of cells to DNA-damaging agents leads to the accumulation of wild-type p53 protein. Furthermore, overexpression of the wild-type p53, mediated by transfection of p53-coding cDNA, induced cells to undergo apoptosis or cell differentiation. In this study we found that the gamma-irradiation that caused the accumulation of wild-type p53 in 70Z/3 pre-B cells induced, in addition to apoptosis, cell differentiation. This was manifested by the expression of the kappa light chain immunoglobulin gene that coincided with the accumulation of cells at the G2 phase. Overexpression of mutant p53 in 70Z/3 cells interferes with both differentiation and accumulation of cells at the G2 phase, as well as with apoptosis, which were induced by gamma-irradiation. Furthermore, the increment in the wild-type p53 protein level following gamma-irradiation was disrupted in the mutant p53 overproducer-derived cell lines. This suggests that mutant p53 may exert a dominant negative effect in all of these activities. Data presented here show that while p53-induced apoptosis is associated with the G1 checkpoint, p53-mediated differentiation, which may be an additional pathway to escape the fixation of genetic errors, may be associated with the G2 growth arrest phase.     

Genetic variation in lipoprotein (a) levels in families enriched for coronary artery disease is determined almost entirely by the apolipoprotein (a) gene locus.

Lipoprotein (a) (Lp[a]) is a cholesterol-rich lipoprotein resembling LDL but also containing a large polypeptide designated apolipoprotein (a) (apo[a]). Its levels are highly variable among individuals and, in a number of studies, are strongly correlated with the risk of coronary artery disease (CAD). In an effort to determine which genes control Lp(a) levels, we have studied 25 multiplex families (comprising 298 members) enriched for CAD. The apo(a) gene was genotyped among the families, using a highly informative pulse-field gel electrophoresis procedure. In addition, polymorphisms of the gene for the other major protein of Lp(a), apolipoprotein B (apoB), were examined. Quantitative sib-pair linkage analysis indicates that apo(a) is the major gene controlling Lp(a) levels in this CAD population (P = .001; 99 sib pairs), whereas the apoB gene demonstrated no significant quantitative linkage effect. We estimate that the apo(a) locus accounts for < or = 98% of variance of Lp(a) serum levels. Approximately 43% of this variation is explained by size polymorphisms within the apo(a) gene. These results indicate that the apo(a) gene is the major determinant of Lp(a) serum levels not only in the general population but also in a high-risk CAD population.     

Estimating the recombination frequency for the MN and the Ss loci

Linkage analysis of 146 informative families for MN and Ss resulted in an estimate of the recombination frequency greater than previously reported. Our total is 7 recombinant children out of 467 individuals, including 1 confirmed recombinant (retested and HLA-compatible) and 6 not verified. The 95% confidence interval of our estimate of recombination is 0.0033-0.0167. Our results are compared with two earlier studies.     

Sample-size calculations in segregation analysis

Segregation analysis, employing nuclear families, is the most frequently used method to evaluate the mode of inheritance of a trait. To our knowledge, there exists no tabular information regarding the sample sizes required of individuals and families needed to perform a significance test of a specific segregation ratio for a predetermined power and significance level. To fill this gap, we have developed sample-size tables based on the asymptotic variance of the maximum likelihood estimate of the segregation ratio and on the normal approximation for two-sided hypothesis testing. Assuming homogeneous sibship size, minimum sample sizes were determined for testing the null hypothesis for the segregation ratio of 1/4 or 1/2 vs. alternative values of .05-.80, for the significance level of .05 and power of .8, for ascertainment probabilities of nearly 0 to 1.0, and sibship sizes 2-7. The results of these calculations indicate a complex interaction of the null and the alternate hypotheses, ascertainment probability, and sibship size in determining the sample size required for simple segregation analysis. The accompanying tables should aid in the appropriate design and cost assessment of future genetic epidemiologic studies.