5' flanking sequences of the murine adenosine deaminase gene direct expression of a reporter gene to specific prenatal and postnatal tissues in transgenic mice.

Adenosine deaminase (ADA), an enzyme of purine metabolism, is highly expressed in four tissues of the mouse: the maternal decidua, the fetal placenta, the keratinizing epithelium of the upper alimentary tract (tongue, esophagus, and forestomach), and the absorptive epithelium of the proximal small intestine. ADA is produced at relatively low levels in all other tissues. To identify genetic elements that direct appropriate prenatal and postnatal expression of the ADA gene, a segment of DNA including the ADA promoter and 6.4 kilobases of the adjacent 5' flanking region was tested for the ability to direct the expression of a reporter gene in transgenic mice. In seven lines of transgenic mice studied, this construct directed high levels of reporter gene expression in the placenta and forestomach and exhibited correct developmental regulation in these tissues. This construct failed to direct significant reporter gene expression to either the maternal decidua or the proximal small intestine. Thus, different gene regulatory elements are required to target high expression to the four tissues characterized by high levels of ADA.     

Regulation of the transmodulated epidermal growth factor receptor by cholera toxin and the protein phosphatase inhibitor okadaic acid.

Addition of tumor promoting phorbol esters, such as phorbol 12-myristate 13-acetate (PMA), to many cell lines results in a decrease of 125I-epidermal growth factor (EGF) binding and increased serine/threonine phosphorylation of the EGF receptor in a process termed transmodulation. It is, however, unclear whether or not receptor phosphorylation is causally related to the inhibition of high affinity EGF binding. We have investigated the significance of phosphorylation/dephosphorylation events in the mechanism of PMA-induced transmodulation using the adenylate cyclase activator cholera toxin and the serine/threonine protein phosphatase inhibitor okadaic acid. In Rat-1 fibroblasts treated at 37 degrees C, PMA induced a rapid decrease in EGF binding which persisted for 3 hours. In contrast, cells exposed to PMA in the presence of cholera toxin exhibited a marked recovery of binding within 60 minutes. The PMA-stimulated decrease in binding correlated with a rapid increase in the phosphorylation state of the EGF receptor. While phosphorylation of the receptor was sustained at an elevated level for at least three hours in cells receiving PMA alone, EGF receptor phosphorylation decreased between 1 and 3 hours in cells treated with PMA and cholera toxin. Furthermore, the cholera toxin-stimulated return of EGF binding was inhibited by treatment with the phosphatase inhibitor okadaic acid. These results suggest that a cholera toxin-activated phosphatase can increase binding capacity of the transmodulated EGF receptor in Rat-1 cells. Cholera toxin treatment elicited a qualitatively similar response in cells transmodulated by platelet-derived growth factor (PDGF). Okadaic acid antagonized the natural return of binding observed in cells stimulated with PDGF alone, indicating that a dephosphorylation event may be required for the recovery of normal EGF binding after receptor transmodulation.     

Male-male behavior and sexual dimorphism of the ear of a zaprochiline tettigoniid (Orthoptera: Tettigoniidae)

The external anatomy of the auditory system of an undescribed zaprochiline tettigoniid (Genus nov. 22 Sp. 1, Australian National Insect Collection, Canberra) shows sexual dimorphism: the male appears to have no auditory spiracle equivalent to that seen in the female. Nocturnally active males aggregate around female required nectar sources in a random manner with regard to each other. There is limited evidence, either from song interaction or from their behavior in the field, that males respond to each other by acoustic cues. Laboratory trials, testing male phonotaxis, showed that movement was random with respect to a target group of caged calling males. In the field, the only signs of agonistic behavior consisted of an increased calling rate when males were close together. Taken together, these data suggest that the male may not preceive sound in the same way as the female.     

Differential response of cycling and noncycling cells to inducers of DNA synthesis and mitosis

The objective of this study was to determine whether cells in G(0) phase are functionally distinct from those in G(1) with regard to their ability to respond to the inducers of DNA synthesis and to retard the cell cycle traverse of the G(2) component after fusion. Synchronized populations of HeLa cells in G(1) and human diploid fibroblasts in G(1) and G(0) phases were separately fused using UV-inactivated Sendai virus with HeLa cells prelabeled with [(3)H]ThdR and synchronized in S or G(2) phases. The kinetics of initiation of DNA synthesis in the nuclei of G(0) and G(1) cells residing in G(0)/S and G(1)/S dikaryons, respectively, were studied as a function of time after fusion. In the G(0)/G(2) and G(1)/G(2) fusions, the rate of entry into mitosis of the heterophasic binucleate cells was monitored in the presence of Colcemid. The effects of protein synthesis inhibition in the G(1) cells, and the UV irradiation of G(0) cells before fusion, on the rate of entry of the G(2) component into mitosis were also studied. The results of this study indicate that DNA synthesis can be induced in G(0)nuclei after fusion between G(0)- and S-phase cells, but G(0) nuclei are much slower than G(1) nuclei in responding to the inducers of DNA synthesis because the chromatin of G(0) cells is more condensed than it is in G(1) cells. A more interesting observation resulting from this study is that G(0) cells is more condensed than it is in G(1) cells. A more interesting observation resulting from this study is that G(0) cells differ from G(1) cells with regard to their effects on the cell cycle progression of the G(2) nucleus into mitosis. This difference between G(0) and G(1) cells appears to depend on certain factors, probably nonhistone proteins, present in G(1) cells but absent in G(0) cells. These factors can be induced in G(0) cells by UV irradiation and inhibited in G(1) cells by cycloheximide treatment.     

Comparative in vitro activity of moxalactam (LY127935), other beta-lactam antibiotics,and aminoglycosides

Moxalactam (LY127935), a novel beta-lactam antibiotic, was compared with semisynthetic penicillins, cephalosporins, and aminoglycosides by the agar dilution method against 5,317 recent clinical isolates of facultative and anaerobic bactria. At 0.5 μg/ml, moxalactam inhibited 90% of all Gram-negative bacilli tested except forPseudomonas aeruginosa (81% inhibited by 32 μg/ml) andAcinetobacter calcoaceticus (88% inhibited by 32 μg/ml). More than 90% ofBacteroides fragilis andStaphylococcus aureus were inhibited by 4 μg/ml and 8 μg/ml, respectively. Moxalactam was at least 16-fold more active by weight than cephalothin, cefamandole, and cefoxitin forEscherichia coli, Klebsiella pneumoniae, andEnterobacter species, and 2- to 4-fold more active than cefoxitin forB. fragilis. Moxalactam was 4-fold less active than cefamandole and cephalothin forS. aureus and 2- to 4-fold less active than piperacillin forP. aeruginosa. Moxalactam was as active or more active than the aminoglycosides for all facultative Gram-negative bacilli except forP. aeruginosa. Moxalactam was inhibitory (minimal inhibitory concentration <16 μg/ml) for 20/27 gentamicin-resistant isolates and 8/13 amikacin-resistant organisms. Moxalactam’s in vitro activity against Gram-negative bacilli is markedly superior to presently available cephalosporins and, except forP. aeruginosa, is comparable to the aminoglycosides.     

Membrane association of a Staphylococcus aureus plasmid in Bacillus subtilis.

The Staphylococcus aureus plasmid pUB110 was found to be enriched in deoxyribonucleic acid-membrane complexes isolated from Bacillus subtilis containing pUB110.     

Characterization of amber and ochre suppressors in Salmonella typhimurium.

Amber and ochre suppressor mutations in Salmonella typhimurium were selected. The amino acid insertions directed by the suppressors were inferred from suppression patterns of Escherichia coli lacI amber mutations. These amber mutations only respond to nonsense suppressors that direct the insertion of particular amino acids. Four Salmonella amber suppressors characterized insert serine, glutamine, tyrosine, and (probably) leucine. Of the three ochre suppressors characterized, two direct the insertion of tyrosine and one directs that of lysine. Of the three amber and two ochre suppressors which have been mapped by phage P22 cotransduction, all are located in the same relative position on the Salmonella map as the analogous E. coli suppressors are on the E. coli map.     

CONTRIBUTION OF POLLEN MORPHOLOGY TO SYSTEMATICS OF COLLOMIA (POLEMONIACEAE)

Pollen morphology of 14 species of Collomia (Polemoniaceae) was examined by light microscopy, and by both scanning and transmission electron microscopy. Four distinct pollen types were observed which are based principally upon 1) shape, number and distribution of apertures, and 2) surface sculpturing: Type 1—zonocolporate with striate ridges; Type 2—zonocolporate with striato-reticulate ridges; Type 3—pantoporate with radiate ridges; Type 4—pantoporate with irregularly reticulate ridges. Evaluation of pollen morphology reveals considerable discrepancy with respect to presently accepted sectional classification. Collomia grandiflora of sect. Collomia has a pollen type similar to that of members of sect. Collomiastrum and is now interpreted as representing an independent evolutionary line derived from the latter section. Collomia diversifolia of sect. Courtoisia has a pollen morphology similar to that of sect. Collomia. whereas C. heterophylla of the same section possesses pollen unique within the genus. This last pollen type shows close similarity to the pollen of members of Polemonium, Gilia, Leptodactylon, and Ipomopsis. Pollen of C. tinctoria and C. tracyi of sect. Collomia are anomalous within Polemoniaceae. No significant difference in exine stratification was discernible among the four pollen types.     

Ascorbate increases the synthesis of procollagen hydroxyproline by cultured fibroblasts from chick embryo tendons without activation or prolyl hydroxylase

An improved procedure was developed to extract prolyl hydroxylase from tendon cells of chick embryos with detergent, and improved assays were developed for both the activity of the enzyme and the amount of enzyme protein. Freshly isolated tendon cells were found to contain approx. 100 μg of enzyme protein per 10⁸ cells and 40–50% of the enzyme protein was active. When the cells were cultured, they were found to contain the same amount of enzyme protein by only 15–20% of the enzyme protein was active. Gel filtration of cell extracts indicated that the active form of prolyl hydroxylase in freshly isolated tendon cells and in cultured tendon cells had the same apparent size and the same activity per μg of immunoreactive protein as enzyme which was shown to be a tetramer. The inactive form was found to have about the same apparent size as subunits of the enzyme.When freshly isolated cells were incubated for 2 h in the presence of 40 μg per ml of ascorbate, there was a slight increase in the rate of hydroxyproline synthesis. In cultured cells, ascorbate at a concentration of 40 μg per ml caused a 2-fold increase in the rate of hydroxyproline synthesis within 30 min. However, ascorbate did not increase the activity of prolyl hydroxylase in extracts from either cell system. Therefore it appears that the influence of ascorbate on synthesis of procollagen hydroxyproline by the cells studied here must be ascribed to a cofactor effect on the hydroxylation reaction similar to that observed with purified enzyme, and it does not involve “activation” of inactive enzyme protein to active enzyme as has been observed in cultures of L-929 and 3T6 mouse fibroblasts.