Characterization of cDNA clones for differentially expressed genes in embryos of dormant and nondormant Avena fatua L. caryopses

The molecular regulation of seed dormancy was investigated using differential display to visualize and isolate cDNAs representing differentially expressed genes during early imbibition of dormant and nondormant Avena fatua L. embryos. Of about 3000 cDNA bands examined, 5 cDNAs hybridized with mRNAs exhibiting dormancy-associated expression patterns during the first 48 h of imbibition, while many more nondormancy-associated cDNAs were observed. Dormancy-associated clone AFD1 hybridized with a 1.5 kb mRNA barely detectable in dry dormant and nondormant embryos that became more abundant in dormant embryos after 24 h of imbibition. Clone AFD2 hybridized with two mRNAs, a 1.3 kb message constitutively expressed in dormant and nondormant embryos and a 0.9 kb message present at higher levels in dormant embryos after 3 h of imbibition. Nondormancy-associated clones AFN1, AFN2 and AFN3 hybridized with 1.5 kb, 1.7 kb and 1.1 kb mRNAs, respectively, that were more abundant in nondormant embryos during imbibition. Expression patterns of some mRNAs in dormant embryos induced to germinate by GA₃ treatment were different than water controls, but were not identical to those observed in nondormant embryos. DNA sequence analysis revealed 76% sequence identity between clone AFN3 and a Citrus sinensis glutathione peroxidase-like cDNA, while significant sequence similarities with known genes were not found for other clones. Southern hybridization analyses showed that all clones represent low (1 to 4) copy number genes.     

Factors affecting levels of genetic diversity in natural populations.

Genetic variability is the clay of evolution, providing the base material on which adaptation and speciation depend. It is often assumed that most interspecific differences in variability are due primarily to population size effects, with bottlenecked populations carrying less variability than those of stable size. However, we show that population bottlenecks are unlikely to be the only factor, even in classic case studies such as the northern elephant seal and the cheetah, where genetic polymorphism is virtually absent. Instead, we suggest that the low levels of variability observed in endangered populations are more likely to result from a combination of publication biases, which tend to inflate the level of variability which is considered ''normal'', and inbreeding effects, which may hasten loss of variability due to drift. To account for species with large population sizes but low variability we advance three hypotheses. First, it is known that certain metapopulation structures can result in effective population sizes far below the census size. Second, there is increasing evidence that heterozygous sites mutate more frequently than equivalent homozygous sites, plausibly because mismatch repair between homologous chromosomes during meiosis provides extra opportunities to mutate. Such a mechanism would undermine the simple relationship between heterozygosity and effective population size. Third, the fact that related species that differ greatly in variability implies that large amounts of variability can be gained or lost rapidly. We argue that such cases are best explained by rapid loss through a genome-wide selective sweep, and suggest a mechanism by which this could come about, based on forced changes to a control gene inducing coevolution in the genes it controls. Our model, based on meiotic drive in mammals, but easily extended to other systems, would tend to facilitate population isolation by generating molecular incompatabilities. Circumstances can even be envisioned in which the process could provide intrinsic impetus to speciation.     

K dependence of the Na-K pump is abnormal in erythrocytes from patients with cystic fibrosis and obligate heterozygotes

The affinity of the Na-K pump for K was significantly (P less than .001) lower in erythrocytes from patients with cystic fibrosis (Km 4.6 +/- 0.35 mM; n = 26) or from heterozygotes (Km 3.9 +/- 0.57 mM; n = 12) than in controls (Km 2.2 +/- 0.10 mM; n = 20). The affinity of the Na-K pump for K was lower in normal erythrocytes than in normal fibroblasts which may explain the variability in the severity of involvement of different organs in cystic fibrosis. We have now shown in human skin fibroblasts and erythrocytes, that the K affinity of the Na-K pump is lower in patients with cystic fibrosis than in controls. Since the abnormality is also present in erythrocytes from heterozygotes who are clinically normal, it is likely that this abnormality is closely related to the genetic defect in cystic fibrosis.     

Protein secretion in phosphate-limited cultures of Bacillus subtilis 168

The secretion of proteins from Bacillus subtilis was studied under physiologically well-defined conditions in continuous cultures at a range of specific growth rates. The kinetics of secretion was analysed by using pulse-chase and immunoprecipitation techniques that allowed both processing and release to be monitored. Growth conditions were selected that were known to lead to significant changes in the anionic polymer composition of the cell wall. Under magnesium limitation only low levels of native proteins were released into the growth medium. In contrast, much higher amounts of released protein were observed under phosphate limitation. Although synthesis of native secretory proteins appeared to be highly regulated, only minor changes in the secretion of heterologous proteins were detected. Comparable kinetics of protein release of cells grown under different conditions indicated similar cell wall permeabilities. The large changes in the amounts of released proteins were not reflected in the production of chaperones and components required for protein secretion. The data suggest that the capacity of the secretion machinery is not a major limiting step in the export of native secretory proteins. Received: 23 September 1997 / Received revision: 10 November 1997 / Accepted: 16 November 1997     

Cloning and sequencing of a plasmid-mediated erythromycin resistance determinant from Staphylococcus xylosus

A 2.3-kb DNA fragment cloned from plasmid pCH200, the largest (52 kb) of four plasmids detected in Staphylococcus xylosus, was found to confer resistance to 14-membered ring macrolides in Bacillus subtilis and Staphylococcus aureus. DNA-sequence analysis of the fragment revealed the presence of an open-reading frame, the deduced product of which was identical to one of the two ATP-binding domains encoded by the macrolide/streptogramin-B-resistance gene msrA of Staphylococcus epidermidis. The observation that a polypeptide homologous to the C-terminus of MsrA is capable of mediating erythromycin resistance in the absence of the N-terminal region is of significance both to the evolution and functional activity of members of the ATP-binding transport super-gene family.     

An order-specific monoclonal antibody to Diptera reveals the impact of alternative prey on spider feeding behavior in a complex food web

Generalist predators have the capacity to restrict pest population growth, especially early in the season before densities increase. However, their polyphagous feeding habits sometimes translate into reduced pest consumption when they target alternative prey. An order-specific monoclonal antibody was developed to examine the strength of trophic connections between Diptera, a major category of non-pest prey, and linyphiid spiders in alfalfa. We report the development and characterization of a monoclonal antibody with order-level specificity to Diptera. This antibody elicited strong absorbance to 22 Diptera from 13 families, no false-positive reactivity to non-dipteran invertebrates, and antigen detection periods following prey consumption that were comparable between spiders. Over 900 field-collected females of the linyphiid spiders Erigone autumnalis and Bathyphantes pallidus were screened for Diptera antigen. Significantly more B. pallidus screened positive for Diptera (40%) compared to E. autumnalis (16%), indicating differential reliance on these prey. In parallel with the collection of spiders for gut-content analysis, prey availability was estimated at web sites. The two spiders exhibited different feeding responses to prey availability. Consumption of Diptera by B. pallidus was strongly correlated with Diptera abundance whilst the availability of other potential prey did not influence predation rates. Conversely, E. autumnalis did not prey upon Diptera in proportion to availability, but increased Collembola activity-density reduced dipteran consumption. Integration of molecular gut-content analysis with precise sampling of prey demonstrated how two closely related linyphiid spiders exhibit different feeding responses to the availability of prey under natural field conditions. Elucidating the feeding preferences of natural enemies is critical to effective incorporation of biological control by generalist predators in the management of agricultural pests.