Nucleotide sequence and expression of the gene for the site-specific integration protein from bacteriophage HP1 of Haemophilus influenzae.

The nucleotide sequence of the leftmost 2,363 base pairs of the HP1 genome, which includes the attachment site (attP) and the integration region, was determined. This sequence contained an open reading frame encoding a 337-residue polypeptide, which is a member of the integrase family of site-specific recombination proteins as judged by sequence comparison. The open reading frame was located immediately adjacent to the att site and was oriented so that initiation of translation would begin distal to the att site and end in its immediate vicinity. Expression of this DNA segment in Escherichia coli provided extracts which promoted site-specific recombination between plasmids containing cloned HP1 attP and Haemophilus influenzae attB sites. This recombination was directional, since no reaction was observed between plasmids containing attR and attL sites. The reaction was stimulated by the accessory protein integration host factor of E. coli. Evidence was also obtained that the integration host factor influenced the levels of HP1 integrase expression. The deduced amino acid sequence of HP1 integrase has remarkable similarity to that deduced for the integrase of coliphage 186.     

Pre- and postjunctional actions of neuropeptide Y and related peptides

The effects of neuropeptide Y (NPY) and related peptide fragments on blood pressure and vagal action at the heart were compared in the anaesthetized rat. A change in vagal action was taken as a measure of presynaptic activity and a change in blood pressure was taken as a measure of postsynaptic activity. NPY, NPY-(13-36), PYY-(13-36), des-Ser22-NPY-(13-36) and a stabilized 13-36 analogue of NPY (ANA NPY) all exerted pressor actions and attenuated vagal action at the heart. The maximum vagal inhibitory or presynaptic action in order of potency was NPY, ANA-NPY, PYY-(13-36) significantly greater than NPY-(13-36), des-Ser22-NPY-(13-36). The order of potency for the half time of this effect was NPY, ANA-NPY significantly longer than PYY-(13-36) and NPY-(13-36), which were significantly longer than des Ser22-NPY-(13-36). For the pressor or postsynaptic effects, NPY increased blood pressure significantly more and for a longer duration than all the 13-36 fragments, which were not demonstrably different in this respect. These results are consistent with the proposal that there are two populations of NPY receptors. The C-terminal flanking peptide of NPY (CPON) and desamido-NPY had no effect on either vagal action at the heart or on blood pressure.     

Guidance of pioneer growth cones: Filopodial contacts and coupling revealed with an antibody to Lucifer Yellow

We are interested in the factors that guide individual neuronal growth cones during embryonic development. We have developed an antibody to the fluorescent dye Lucifer Yellow. We use the antibody here to examine the specific filopodial contacts and dye coupling by the first growth cones in the grasshopper embryo that navigate in an axonless environment. We have studied the distribution and apparent selective adhesion of the filopodia from these pioneering growth cones in the central nervous system and periphery. Our results suggest that selective filopodial adhesion to specific “landmark” cells may play an important role in the guidance of pioneer growth cones.     

Variation within teosinte. II. numerical analysis of chromosome knob data

Principal components analysis of 46 chromosome knob positions and B chromosomes for 61 accessions of annual teosinte (in 12 groups) and 207 accessions of maize (in 87 groups) produced results which did not entirely agree with previous groupings. Teosintes of northwest and southeast Guatemala (races Huehuetenango and Guatemala) were widely separated from maize and from all other teosintes; however, teosintes of east Mexico-Distrito Federal (race Chalco) were also widely separated from maize and other teosintes. Teosintes of south Chihuahua (race Nobogame) grouped separately from other teosintes, suggesting that race Nobogame is not simply a northern extreme of race Central Plateau. Teosintes of east Michoacan and west Mexico (recently grouped into the Balsas race) grouped together but separately from other teosintes. The latter teosintes may merit special designation in a racial taxonomic system. Only the Central Plateau teosintes consistently overlapped with maize OTU's for the first few principal components. Even in that case there was no consistent association between the Central Plateau teosintes and any specific race of maize.     

Juvenile hormone esterases in diapause and nondiapause larvae of the European corn borer, Ostrinia nubilalis

Haemolymph levels of juvenile hormone esterase, 1-naphthyl acetate esterase, and juvenile hormone were measured in synchronously staged diapause and nondiapause larvae of the European corn borer, Ostrinia nubilalis. Juvenile hormone esterase levels were monitored using juvenile hormone I as a substrate while juvenile hormone titres were measured with the Galleria bioassay. Haemolymph of nondiapause larvae showed two peaks of juvenile hormone hydrolytic activity: one near the end of the feeding phase and a smaller one just prior to pupal ecdysis. These peaks of enzyme activity correlated well with the low levels of haemolymph juvenile hormone. Juvenile hormone titres were high early in the stadium then showed a second peak during the prepupal stage coinciding with low esterase activity. Diapause haemolymph had peak juvenile hormone esterase activity nearly 4 times the nondiapause level, reaching a peak near the end of the feeding phase. Diapause-destined larvae retained high juvenile hormone titres even during the rise of the high esterase levels. 1-naphthyl acetate esterase levels did not correlate with the juvenile hormone esterase levels in either the diapause or nondiapause haemolymph. High levels of 1-naphthyl acetate esterase activity were associated with moulting periods.     

Isolation of the trypsin inhibitors in Ascaris lumbricoides var. suum using affinity chromatography

A method for isolating three water-soluble trypsin inhibitors from Ascaris lumbricoides var. suum by affinity chromatography is described. The trypsin inhibitors captured by affinity chromatography are resolved into three species by chromatography on CM-Sephadex at pH 8.1. The inhibitors are named in the order that they are released from the CM-Sephadex column. Ascaris Trypsin Inhibitor 1 is the same as inhibitor CM-1 described by [3.] and inhibitor Peak I of U. Kucich and [4.]. Ascaris Trypsin Inhibitor 2 is the inhibitor described by [2.] and inhibitor CM-2 of [3.]. Ascaris Trypsin Inhibitor 3 is the same as inhibitor Peak II of [4.]. Ascaris Trypsin Inhibitor 1 is 80%, Ascaris Trypsin Inhibitor 2 is 8%, and Ascaris Trypsin Inhibitor 3 is 12% of the water-soluble trypsin inhibitors present in Ascaris. With this procedure all of the Ascaris trypsin inhibitors can be isolated in a few days. This shortens the exposure of personnel to crude extracts of Ascaris and diminishes the biological hazard of working with them. Frequent exposure to Ascaris extracts may evoke an anaphylactic response in personnel.     

Electron-microscopic and morphometric study of vesiculation in the epithelial cell layer of the toad urinary bladder

Summary The sequence of events in the establishment of a keratinised epidermis has been related to age and crown rump length in fetal rats. Differentiation of the epidermis occurs under cover of the periderm throughout gestation. Ten stages are defined between 12d intra-uterine life and birth. Membrane-coating granules (MCGs) appeared at 18d (Stage 4) after the appearance of tonofilaments but before the appearance of the first fetal keratohyaline granules (KHGs) at 19d (Stage 5). Measurement of the position of MCGs within the cells showed a less marked concentration near the superficial border of the cells than that found at later stages. As KHGs formed, exocytosis of MCGs occurred into the intercellular space immediately deep to the periderm. It is suggested that after 19d (Stage 5) the periderm serves to retain the contents of the MCGs in this space so providing the permeability barrier and that prior to that stage the periderm itself probably provides the permeability barrier of fetal skin.     

Molecular cloning of the avian myelocytomatosis virus genome and recovery of infectious virus by transfection of chicken cells.

The avian retrovirus myelocytomatosis virus 19 (MCV) possesses an interesting diversity of oncogenic potentials, but the virus has proven difficult to study because of its inability to replicate without the assistance of a helper virus. We have therefore isolated and amplified the genome of MCV by molecular cloning in a procaryotic vector. The topography of the cloned DNA was explored by the use of restriction endonucleases and radioactive complementary DNAs representing specific domains in avian retrovirus genomes. The cloned DNA appeared to be an authentic representation of the MCV genome: the size and genetic topography of the DNA were comparable to those of MCV, and transfection of the cloned DNA into chicken cells (in company with the DNA of a suitable helper virus) gave rise to virus with the genome and transforming potentials of MCV. The availability of cloned MCV DNA should facilitate a variety of genetic and biochemical manipulations directed at elucidating the mechanism of oncogenesis by MCV.     

Lysis of Streptococcus mutans by hen egg white lysozyme and inorganic sodium salts.

Streptococcus mutans BHT was grown in a synthetic medium containing radioactive thymidine to monitor deoxyribonucleic acid release. Kinetic experiments demonstrated that although lysozyme alone could not liberate deoxyribonucleic acid, cellular deoxyribonucleic acid was liberated from lysozyme-treated cells by addition of low concentrations of inorganic sodium salts. When the salts were tested for their ability to dislodge cell-bound tritiated lysozyme, the extent of the initial release of enzyme by individual anions correlated with the anion potency for deoxyribonucleic acid liberation (SCN- greater than ClO4- greater than I- greater than Br- greater than NO3- greater than Cl- greater than F-), although the total amount of lysozyme dislodged did not correspond directly with cell lysis. Differences in the effectiveness of anions (SCN-, HCO3-, Cl- and F-) in potentiating cell lysis could be enhanced or minimized by varying the lysozyme, anion, and bacterial cell concentrations. As the anion concentration was increased for each enzyme concentration and cell concentration, the lysis increased, in some cases markedly, until maximum levels of released deoxyribonucleic acid were attained. The maximum levels of lysis of SCN- and HCO3- were similar and were greater than those for Cl- and F-. In addition, the maximum levels were observed to increase for each of the anions as the concentration of lysozyme increased.