A primer ribonucleic acid for initiation of in vitro Rous sarcarcoma virus deoxyribonucleic acid synthesis.

The nucleotide sequence of an RNA primer molecule for initiation of Rous sarcoma virus DNA synthesis in vitro has been determined. The sequence can be drawn in a cloverleaf structure typical of tRNAs with an anticodon for tryptophan. Aminoacylation of the molecule confirms that it is tRNA-Trp. The same sequence and aminoacylation results are obtained regardless of whether the RNA is isolated from virions or from cells of chickens, the natural host for this virus. It is the only species of tRNA-Trp that is dectected in chicked cell tRNA.     

Standards for biodiversity: a proposal based on biodiversity standards for forest plantations

The 1992 Convention on Biological Diversity has not only focused attention on the concept of biodiversity it has also set expectations that the signatory nations will establish objectives. It is suggested here that the use of biodiversity standards could have a useful role in quantifying these objectives. The use of biodiversity standards is explored in connection with the levels of biodiversity In plantation forests. Forest plantations have been used for the basis of this investigation because such plantations could be used as large-scale experiments in which the use of standards, and the ecological monitoring of these standards, could be researched. The impact of intensive forestry practices on wildlife has often been the cause of concern and indeed controversy. Recently there has been a move towards the development of multipurpose forest plantations where management for recreation and wildlife takes place alongside management for timber. Some forest authorities are actively looking at ways of restoring levels of biodiversity in plantation forests. We suggest that if levels of biodiversity are to be restored, then forest managers must have something to aim for; that is, there must be a standard. The adoption of standards could provide a means whereby forest managers can meet biodiversity objectives. Standards may also enable managers to determine the effectiveness of those management practices which are intended to increase levels of biodiversity. We suggest a conceptual plan which combines a monitoring procedure with biodiversity objectives.     

Mylohyoid and jugular foramen bridging in pre-Columbian Chileans

The occurrence of mylohyoid groove bridging and jugular foramen bridging was determined in a population of adult Pre-Columbian Chileans. Two hundred forty-one crania (110 males, 131 females) were examined for jugular foramen bridging and 464 mandibles (252 males and 212 females) were observed for mylohyoid groove bridging. The overall incidence of jugular foramen bridging was 14.94%, with females showing a higher occurrence (18.32%) than males (10.91%); however, no statistical significance was found between the sexes. The right side showed a greater incidence than the left, but again no statistical difference was noted and the occurrence of this trait was found to be associated between the sides. This frequency of jugular foramen bridging is higher than that found in other studies and approaches that reported in North American Eskimo populations. In the case of mylohyoid groove bridging, the incidence was 4.09% with no difference found between the sexes. This frequency of occurrence is less than the values reported previously for the other American populations of Mongoloid ancestry including the Pre-Columbian Peruvian population. The importance of nonmetric skeletal traits for defining population groups and their usefulness as anthropological tools is discussed. It is concluded that hard tissue variations such as bony bridging may be reliable markers to aid in discretely identifying population groups, but their importance can be significantly enhanced by coupling as many traits as possible in future studies.     

Expression and sorting of rat plasma kallikrein in POMC-producing AtT-20 cells.

A vaccinia virus (VV) vector was used to express rat plasma kallikrein (rPK) in the constitutively secreting cells, BSC-40, and in the endocrine regulated cells, AtT-20. Using a specific rPK antibody and a fluorogenic substrate, Phe-Phe-Arg-AMC, we demonstrated that in both cell lines VV infections resulted in the synthesis of an immunoreactive enzyme predominantly present as a zymogen which can be activated with trypsin. Stimulation of VV:rPK-infected AtT-20 cells with either 5mM 8-bromo-cAMP or 56 mM KCl resulted in a different pattern of rPK and ACTH secretion, strongly suggesting that rPK follows the constitutive secretory pathway. Finally, the 10% rPK activity found within AtT-20 cell extracts had no effect on pro-opiomelanocortin (POMC) processing either intracellularly or extracellularly. The above data show that the biosynthetic machinery of both cell lines analyzed does not allow the efficient activation of plasma prekallikrein. Finally, despite the PK's demonstrated ability to cleave various hormone precursors in vitro at pairs of basic residues, in vivo, we did not obtain evidence that this hepatic enzyme can also act as an intracellular pro-protein processing enzyme.     

Rapid detection of antigen binding by antibody fragments expressed in the periplasm of Escherichia coli.

Bacterial expression systems can greatly facilitate protein engineering of antibodies. We have developed a system for high-level expression of antibodies, antibody fragments, or hybrid antibodies with novel effector functions in the periplasm of Escherichia coli. From 5 ml of cells, a simple extraction yields sufficient material for SDS-gel electrophoresis, detection and characterization of hapten binding. To demonstrate our system, heavy-chain variable regions and lambda 1 light chains of a mouse anti-NP antibody were synthesized as hybrid proteins with a bacterial signal peptide (Omp F). Each chain is secreted into the periplasm where processing (cleavage of the signal peptide), folding and heterodimer association take place. Periplasmic proteins are released by cold osmotic shock, and hapten-binding activity is easily detected without further manipulation. The ease of genetic engineering in this system will facilitate the production of immunoglobulin derivatives designed for specific applications, and expression of these molecules in a native state will allow the rapid screening of combinatorial libraries and the results of mutagenesis.     

Depth-dependent fluorescent quenching of a tryptophan residue located at defined positions on a rigid 21-peptide helix in liposomes

Five lipophilic 21-peptide analogs of the potential-dependent pore-former, alamethicin, were synthesized bearing tryptophan residues at the position 1, 6, 11, 16 and 21 on a long, conformationally rigid, alpha-helix. The alpha-helical conformation was induced and stabilized using the sequential oligomers (Ala-Aib-Ala-Aib-Ala)n as analyzed by CD and NMR. The partitioning of the N-t-butoxycarbonyl 21-peptide methyl esters and the N-terminally deprotected alpha-helices was followed by fluorescence enhancement in phospholipid bilayer vesicles. Quenching experiments were performed by titrating with n-doxyl stearic acids bearing the nitroxide label at positions 5, 7, 10, 12 and 16. This well-defined system revealed that the N- and C-terminal tryptophan residues become situated in the hydrophilic region. Tryptophan at position 11 was found in the lipophilic core, whereas the tryptophan at positions 6 and 16 were localized at intermediate depths of the lipid membrane. Therefore, the helices span the lipid bilayer with their long axis normal to the membrane surface.     

Contributions of vasopressin and other pressor systems to DOC-salt hypertension in rats

The roles of arginine vasopressin (AVP), the sympathetic nervous system, and the renin-angiotensin system in maintaining elevated blood pressure in established DOC-salt hypertension in rats were studied by injection of specific antagonists of these systems. The specific AVP antagonist dPVDAVP decreased blood pressure by 19 +/- 3 mm Hg in hypertensive rats and 6 +/- 2 mm Hg in control rats. In a different group of rats ganglionic blockade with chlorisondamine also caused a greater decrease in blood pressure in DOC-salt rats compared to controls (99 +/- 6 vs 58 +/- 4 mm Hg, respectively). In rats with autonomic ganglia blocked subsequent vasopressin antagonism decreased blood pressure 29 +/- 4 mm Hg in DOC-salt rats and 14 +/- 2 mm Hg in control rats. Converting enzyme inhibition with captopril in rats with autonomic ganglia blocked caused a lesser decrease in blood pressure in DOC-salt rats than in controls (8 +/- 2 vs 14 +/- 2 mm Hg, respectively). These results indicate that both AVP and the sympathetic nervous system contribute to the maintenance of DOC-salt hypertension. The renin-angiotension system appears to be relatively less important.