DNA sequence organization in the genomes of five marine invertebrates

The arrangement of repetitive and non-repetitive sequence was studied in the genomic DNA of the oyster (Crassostrea virginica), the surf clam (Spisula solidissima), the horseshoe crab (Limulus polyphemus), a nemertean worm (Cerebratulus lacteus) and a jelly-fish (Aurelia aurita). Except for the jellyfish these animals belong to the protostomial branch of animal evolution, for which little information regarding DNA sequence organization has previously been available. The reassociation kinetics of short (250-300 nucleotide) and long (2,000-3,000 nucleotide) DNA fragments was studied by the hydroxyapatite method. It was shown that in each case a major fraction of the DNA consists of single copy sequences less than about 3,000 nucleotides in length, interspersed with short repetitive sequences. The lengths of the repetitive sequences were estimated by optical hyperchromicity and S1 nuclease measurements made on renaturation products. All the genomes studied include a prominent fraction of interspersed repetitive sequences about 300 nucleotides in length, as well as longer repetitive sequence regions.     

Binding of glucocorticoid receptors to DNA.

DNA has been implicated as the nuclear acceptor for receptor-glucocorticoid complexes. The present study concerns the interaction of these complexes, isolated from cultured rat hepatoma cells, with purified DNA. This association is rapid, reaching a maximum within a few minutes at 0 degrees, whereas dissociation requires several hours. DNA binds neither free glucocorticoids nor those complexed with transcortin or cytosol proteins different from the receptor. Receptors which are not complexed by steroid have little or no affinity for DNA. "Activation," necessary for the binding of receptor-steroid complexes to isolated nuclei, also enhances DNA binding. The capacity of DNA for binding receptor-steroid complexes is large; saturation was not observed at the complex concentrations studied, using either crude or partially purified receptor preparations. The association of complexes with DNA is inhibited by divalent cations, at increasing ionic strengths, and by mercurial reagents. Complexes bind equally well to bacterial, bacteriophage, or rat DNA; however, there was either no or substantially reduced binding by bacterial 23 S rRNA. The binding of complexes to native DNA is roughly 3-fold greater than to denatured DNA. These characteristics are consistent with the possibility that DNA is the nuclear acceptor for receptor-glucocorticoid complexes; however, the actual composition of the acceptor sites remains unknown.     

Organization of mesosomes in fixed and unfixed cells.

After the addition of glutaraldehyde (GA) to cells incubated at 3 or 37 degrees C, mesosomes were observed with increasing frequencies in freeze fractures of cells. These increases were related to the kinetics with which GA cross-linked adjacent amino acids. Upon the addition of GA, mesosomes were first observed in the periphery of freeze-fractured cells usually attached to septal membranes. However, the time, while the septal attachment sites were maintained, the "bodies" of the mesosomes were observed to move toward the center of the cytoplasm. This centralization process was much more rapid at 37 than at 3 degrees C. It is hypothesized that upon fixation, or receipt of some physical insult, mesosome precursors found in undisturbed cells undergo a change in state that results in their visibility in freeze fractures.     

Nuclear components responsible for the retention of steroid--receptor complexes, especially from the standpoint of the specifcity of hormonal responses.

1. By covalently linking nuclear components from hormone-sensitive cells to Sepharose 2B, it is possible to investigate the interaction between nuclear components and cytoplasmic receptor-steroid complexes by affinity chromatography. 2. Many factors are implicated in the specifity of nuclear-cytoplasmic interactions, including the nature of the nuclear components, the presence of the cytoplasmic receptor protein and the provision of the appropriate steroid ligand. 3. Two distinct sets of binding sites are present in nuclear extracts immobilized to Sepharose 2B, namely a small number of specific high-affinity sites and a larger number of non-specific low affinity-sites. 4. Considerable evidence supports the importance of the high-affinity binding sites in the manifestation of hormonal specificity in different tissues. Although the study has centred largely on androgenresponsive systems, the findings are germane to cytoplasmic-nuclear interactions in general. 5. The high-affinity or acceptor sites in nuclear extracts reside in the basic but non-histone protein fraction. 6. Hormonal specificity is seemingly maintained by both the cytoplasmic and nuclear components, and the results are discussed in the context of the mechanism of action of steroid hormones.     

Influence of genetic predisposition to diabetes and obesity on mitochondrial function

Inbred mice with the mutation diabetes C57BL/KsJ db+/db+ and the mutation obese C57BL/6J ob/ob displayed a total liver mitochondrial capacity to oxidize glutamate or succinate which was approximately eight times greater than the capacity of the C57BL/6J +/+ control mice. This increase in oxidation capacity was estimated by multiplying the observed twofold increase in each of the following components: total liver weight, the mitochondrial protein content per gram of liver, and glutamate or succinate respiration activity per milligram of liver mitochondrial protein. No significant difference in liver mitochondrial function and capacity for oxidation was observed between db+/db+ and ob/ob mutants, which indicated that these results may be primarily mediated by the genetic factors responsible for obesity and hyperphagia in these mutants, and not by the genetic traits associated with diabetes. These findings may provide a biochemical foundation in support of the thrifty gene hypothesis.     

Genetic map of the opp (Oligopeptide permease) locus of Salmonella typhimurium.

The uptake of peptides by Salmonella typhimurium is mediated by three apparently independent transport systems. One of these systems, the oligopeptide permease, is encoded by a genetic locus (opp) which has been mapped at 34 min on the S. typhimurium chromosomal map. We accurately mapped the location of opp by cotransduction frequencies and by deletion analysis and show that the gene order for this region of the chromosome is cysB-trp-tonB-opp-galU-tdk. All opp mutants, independently isolated by a variety of means, mapped at this one locus, between tonB and galU. Spontaneous and transposon Tn10-generated deletions were used to construct a fine-structure genetic map of opp. Evidence is presented which indicates that opp covers a 5- to 6-kb segment of DNA and is therefore likely to consist of more than one gene.     

Complement driven innate immune response to malaria: fuelling severe malarial diseases

Severe malaria remains a major cause of global mortality. The innate immune response to infection is a key determinant of malaria severity and outcome. The complement system plays a key role in initiating and augmenting innate immune responses, including inflammation, endothelial activation, opsonization and coagulation, processes which have been implicated in malaria pathogenesis. In this review, we discuss the evidence supporting a role for excessive complement activation in the pathogenesis of severe malaria.     

A sequence-specific function for the N-terminal signal-like sequence of the TonB protein

TonB is a proline-rich protein which provides a functional link between the inner and outer membranes of Gram-negative bacteria. TonB is anchored to the inner membrane via an N-terminal signal-like sequence and spans the periplasm, interacting with transport receptors in the outer membrane. We have investigated the role of the N-terminal signal-like peptide in TonB function. Replacement of the N-terminal sequence with heterologous sequences indicates that it has at least three distinct rotes in TonB function: (i) to facilitate translocation of TonB across the cytoplasmic membrane; (ii) to anchor TonB to the cytoplasmic membrane; (iii) a sequence-specific functional interaction with the ExbBD proteins.