Heteropolypeptides from hydrogen cyanide and water? Solid state15N NMR investigations

Cross-polarization magic-angle spinning¹⁵NMR spectra have been used to determine the composition of hydrogen cyanide polymers both before and after treatment with water. The unambiguous presence of secondary amide groups (as in peptide links) has been established by double-cross-polarization studies on the polymers synthesized from equimolar amounts of H¹³CN and HC¹⁵N. The NMR results are consistent with the hypothesis that the original heteropolypeptides on Earth were synthesized directly from hydrogen cyanide and water without the intervening formation of -amino acids.     

Hyperpolarizing potentials in guinea pig hippocampal CA3 neurons

There is a bewildering variety of hyperpolarizing potentials which control activity in hippocampal pyramidal cells. These include an inhibitory postsynaptic potential (IPSP) with early and late components, voltage- and calcium-dependent potassium conductances, a voltage-dependent potassium conductance modulated by muscarinic agents (the M-current), and a complex and poorly understood afterhyperpolarization following epileptiform bursts. In hippocampal CA3 pyramidal cells, mossy fiber stimulation elicits an IPSP which is made up of two readily separable components. Using the in vitro slice preparation, we investigated the underlying ionic basis of these IPSP components and compared them to other hyperpolarizing potentials characteristic of the CA3 neurons. Intracellular recordings were obtained and then tissue was exposed to bathing medium low in chloride concentration or high in potassium concentration; the ion "blockers" EGTA (intracellular); tetraethylammonium (TEA) (intra- and extracellular), and barium and cobalt (extracellular); and the gamma-aminobutyric acid (GABA)/chloride antagonists penicillin, bicuculline and picrotoxin.     

Nonsense RNA: a tool for specifically inhibiting the expression of a gene in vivo

We describe a general technique to inhibit gene expression in eukaryotic cells. The gene we chose to inhibit was the E. coli LacZ gene (encoding beta-galactosidase), which has previously been cloned into a eukaryotic expression vector [1]. This plasmid is called pCH110. We constructed a variant of pCH110 in which we flipped a 2566 base pair 5' fragment of the LacZ gene into the antiparallel orientation. The plasmid containing this mutated LacZ gene is called pNSLacZ (NS signifies non-sense coding sequence). When equal amounts of pCH110 and pNSLacZ are co-transfected into 3T6 mouse fibroblasts, the beta-galactosidase activity is decreased by approximately a factor of ten. Increasing the ratio of pNSLacZ to pCH110 above 1:1 does not appreciably increase the level of inhibition. Next, we prove the specificity of the inhibition by adding a third gene to the transfection mixture. For this purpose, we used pSVneo beta, a plasmid which expresses a phosphotransferase. We found that even when the beta-galactosidase activity was diminished by a factor of 10, the phosphotransferase activity was unaffected. Therefore, we have demonstrated that: the presence of an antiparallel copy of the LacZ gene results in a significant and specific diminution of the LacZ gene's expression; only a fraction of the LacZ gene needs to be in the antiparallel orientation in order to observe this effect. These results suggest that this technique can serve as a tool to decrease the level of gene expression in order to study the function of specific genes, or as a therapeutic manoeuvre in the treatment of disorders of abnormal gene expression.     

Chemotaxonomic relationships within the order charadriiformes

The uropygial gland secretions of Charadriiformes species (Thinocorus rumicivorus, Gallinago gallinago, Scolopax rusticola) were analysed and the quantitative compositions compared with those of other Charadriiformes and Lariformes species. The results are discussed from a chemotaxonomic viewpoint and evidence for a close relationship between Thinocorus and other Charadriiformes is given.     

Interaction of peanut agglutinin,a lectin specific for nonreducing terminal d-galactosyl residues,with embryonal carcinoma cells

Peanut agglutinin (PNA), a lectin specific for terminal d-galactosyl residues, was found to react with embryonal carcinoma cells, but not with their differentiated derivatives. Receptors for PNA were detectable at the surface of all cells of the quasinullipotent F9 line and on only 50% of the multipotent PCC3/A/1 line. The fraction of the PCC3/A/1 population which expresses the F9 antigen was found to be included in the subpopulation carrying the PNA receptors. PNA⁺ and PNA^? subpopulations of PCC3/A/1 were separated by a PNA-mediated reversible agglutination of PNA⁺ cells with rabbit erythrocytes. These subpopulations were essentially F9⁺ and F9^?, respectively.     

PCC4, a new cell surface antigen common to multipotential embryonal carcinoma cells, spermatozoa, and mouse early embryos

Cell surface antigens of a multipotential embryonal carcinoma (EC) line, PCC4, have been investigated. As do other EC cells, these cells express the F9 antigen but not the H-2 or Ia antigens. In addition, these cells express another antigen called PCC4. This antigen is present on the multipotential EC cells tested, on spermatozoa, and on the inner cell mass of newly implanted blastocyst.     

Anatomy of Herpes Simplex Virus DNA. VIII. Properties of the Replicating DNA

This paper concerns the properties of herpes simplex virus 1 DNA replicating in HEp-2 and human embryonic lung cells. The results were as follows. (i) Only a small fraction of input viral DNA entered the replicative pool. The bulk of the input viral DNA cosedimented with marker viral DNA and did not appear to be degraded or dissociated into L and S components. (ii) Nascent DNA sedimented faster and banded at a higher density than that of mature viral DNA extracted from virions. Pulse-chase experiments indicated that nascent DNA acquires the sedimentation rate and buoyant density of viral DNA within 30 to 40 min after its synthesis. (iii) Electron microscopic studies indicated that the DNA extracted from cells replicating viral DNA and banding at the density of viral DNA contained: (a) linear, full-size molecules with internal gaps and single-stranded regions at termini; (b) molecules with lariats, consisting of a linear segment up to 2x the size of mature DNA and a ring ranging from 0.5 x 10(6) to 100 x 10(6) in molecular weight, showing continuous and discontinuous forks; (c) circular, double-stranded molecules, both full-size and multiples of 18 x 10(6) in molecular weight, but without forks or loops; (d) molecules showing "eye" and "D" loops at or near one end of the DNA; (e) large, tangled masses of DNA, similar to those observed for T4 and pseudorabies virus replicating DNAs, containing loops and continuous and discontinuous forks. The electron micrographs are consistent with the hypothesis that the single-stranded ends on the DNA anneal to form a hairpin, that the DNA synthesis is initiated at or near that end and proceeds bidirectionally to form a lariat, and that resulting progeny derived by semiconservative replication are "head-to-head" and "tail-to-tail" dimers.     

Variation of mitochondrial volume fraction along multiply innervated fibers in rabbit extraocular muscle

Mitochondrial volume fraction was compared among three regions along the length of six multiply innervated fibers (MIFs) in the orbital surface layer of rabbit superior rectus. These MIFs are of about 5 μm diameter toward the middle of their length, and of about 15 μm diameter toward their proximal and distal ends. The region of highest volume fraction (26%) was located toward the proximal end of their segment of minimal diameter, in apparent association with endplate-like nerve junctions. The region of lowest volume fraction (8%) was located at their distal segment of maximal diameter. The region toward the distal end of their segment of minimal diameter displayed an intermediate volume fraction (15%). These mitochondrial volume fractions were further analyzed in terms of the relative contributions of the I-band, the A-band, and the subsarcolemmal mitochondrial clusters. Comparable changes in mitochondrial content occur in both the I-band and A-band: in the fibers' distal segment of maximal diameter, however, the mitochondrial volume fraction in the A-band (5%) is lower than in the I-band (11%). These modifications of mitochondrial content along the fibers' length occur irrespective of the contributions of the subsarcolemmal mitochondrial clusters.     

A monoclonal IgG4 (lambda) with factor V inhibitory activity.

A monoclonal IgG4 (lambda) with inhibitory activity to human coagulation factor V was isolated from the serum of a patient with a fatal hemorrhagic diathesis by using a combination of DE-52 ion exchange chromatography and isoelectric focusing techniques. Using the criteria for defining a monoclonal immunoglobulin of restricted mobility on protein electrophoresis, immunoelectrophoresis, and isoelectric focusing, as well as neutralization with class, subclass, and light chain type antisera, we are the first to demonstrate a factor V inhibitor as a monoclonal IgG4 (lambda) detectable in serum or plasma.     

Stereospecific effects of opiate antagonists on superficial and deep nociception and on motor activity suggest involvement of endorphins on different opioid receptors.

It is known that various opiate antagonists enhance stereospecifically reactions to superficial nociceptive stimuli (e.g. in the hot plate test) suggesting the involvement of endogenous ligands in these reactions. In mice and rats the writhing responses to deep nociceptive stimuli (intraperitoneal test) were also enhanced stereospecifically by (-) naloxone, Mr 2266 and GPA 2163 but some other antagonists (naltrexone, levallorphan, diprenorphine) were inactive probably as a consequence of interfering agonist (antinociceptive) properties. An another antagonist, (-) Win 44441 suggested to bind principally with κ receptors did not enhance either superficial or deep nociception indicating that the former antagonists are probably interfering with endorphins at the level of μ receptors. The motor reaction of mice to a novel environment was stereospecifically depressed by opioid antagonists including (-) Win 44441 suggesting an involvement of endorphins at the level of κ receptors ; Mr 2266 and GPA 2163 were ineffective in this test and hyperalgesic in the two antinociceptive tests ; they might be relatively pure μ antagonists.