Mapping and characterization of a novel human myc-like (MYCLK1) sequence.

The myc family of proto-oncogenes consists of several members that possess regions of sequence homology and some have known similarities in structure and function. We have isolated an 8.8-kb EcoRI fragment from a human genomic library by hybridization to a 28-base oligonucleotide probe derived from a region of the second exon of MYC, which is highly conserved in the myc gene family. Sequence analysis of this myc-like (MYCLK1) DNA fragment has revealed the existence of a region with 85% homology to the 28-base oligonucleotide probe. An open reading frame of 207 nucleotides containing the region of homology was found. We have mapped MYCLK1 to human chromosome 7 at band p15 by chromosome in situ hybridization; this site is distinct from the map location of previously characterized myc genes. Whether MYCLK1 represents a new functional member of the myc family of proto-oncogenes remains to be determined.     

Insulin-stimulated hexose transport and glucose oxidation in rat adipocytes is inhibited by sphingosine at a step after insulin binding

Spingosine, a naturally occurring inhibitor of protein kinase C, has recently been shown to have potent bioregulatory effects on a variety of cellular processes involving signal transduction mechanisms. In the present studies, we have investigated its effects on activation by insulin of hexose transport and glucose oxidation in isolated rat adipocytes. Preincubation of cells with this long-chain base blocked both the marked activation of these processes by insulin and the smaller activation by phorbol myristate acetate. Inhibition of both insulin and phorbol 12-myristate 13-acetate activation showed the same sphingosine concentration dependence, suggesting a common locus of action. The effectiveness of sphingosine was inversely proportional to the lipid content in the incubation (which was a function of both the age of the animal and the number of cells used) presumably due to dilution of the lipophilic long-chain base into the cellular triglycerides. Sphingosine did not affect either insulin binding to its receptor or the half-maximal concentration of the hormone required to activate hexose transport, but reduced the maximal responses. Thus, the inhibition was at a step distal to the binding of insulin to its receptor. Basal transport activity was not inhibited, suggesting a locus of action prior to the glucose transporter. The inhibitor was also effective when added following activation by insulin of hexose transport and resulted in a rapid reversal of activation (t 1/2 for inhibition was 2-4 min.). Sphingosine and its analogs showed a parallel potency for inhibition both of isolated protein kinase C and of insulin activation in adipocytes, consistent with an essential role for protein kinase C in the activation of hexose transport by insulin.     

The osmotic concentration of parietal muscle of the hagfish Myxine glutinosa L., including estimates of free and bound constituents

The principal osmotic constituents of plasma and of muscle before and after ultracentrifugation have been determined. By analysing the muscle fluid and centrifuged muscle and determining their extracellular fluid (inulin space), ion-binding in the cells was estimated at Na 26%, K 0.3%, Ca 93%, Mg 24%, Cl 21% and P 10%. Muscle fluid was 4.9% (2.7-7.4%) hyperosmotic to plasma. This is discussed in relation to calculated osmolality of muscle and plasma.     

Foraging interactions between Wandering Albatrosses Diomedea exulans breeding on Marion Island and long-line fisheries in the southern Indian Ocean

Wandering Albatrosses Diomedea exulans are frequently killed when they attempt to scavenge baited hooks deployed by long-line fishing vessels. We studied the foraging ecology of Wandering Albatrosses breeding on Marion Island in order to assess the scale of interactions with known long-line fishing fleets. During incubation and late chick-rearing, birds foraged further away from the island, in warmer waters, and showed high spatial overlap with areas of intense tuna Thunnus spp. long-line fishing. During early chick-rearing, birds made shorter foraging trips and showed higher spatial overlap with the local Patagonian Toothfish Dissostichus eleginoides long-line fishery. Tracks of birds returning with offal from the Toothfish fishery showed a strong association with positions at which Toothfish long-lines were set and most diet samples taken during this stage contained fishery-related items. Independent of these seasonal differences, females foraged further from the islands and in warmer waters than males. Consequently, female distribution overlapped more with tuna long-line fisheries, whereas males interacted more with the Toothfish long-line fishery. These factors could lead to differences in the survival probabilities of males and females. Non-breeding birds foraged in warmer waters and showed the highest spatial overlap with tuna long-line fishing areas. The foraging distribution of Marion Island birds showed most spatial overlap with birds from the neighbouring Crozet Islands during the late chick-rearing and non-breeding periods. These areas of foraging overlap also coincided with areas of intense tuna long-line fishing south of Africa. As the population trends of Wandering Albatrosses at these two localities are very similar, it is possible that incidental mortality during the periods when these two populations show the highest spatial overlap could be driving these trends.     

A retrospective study of 932 second trimester terminations using gemeprost (16,16 dimethyl-trans delta 2 PGE1 methyl ester).

A retrospective study of 932 second trimester terminations between 12-27 weeks gestation was carried out to determine the efficacy of gemeprost for second trimester termination. A single course of 5 x 1 mg gemeprost pessaries was administered every three hours. If abortion had not occurred after the first course of pessaries, a further course of 5 x 1 mg pessaries was administered. Intravenous oxytocin was administered after 36 hours if abortion had not occurred. Eighty per cent and ninety five per cent of patients aborted within 24 and 48 hours respectively. Of the remaining 5 per cent of women, 3 per cent aborted with escalating doses of oxytocin. In the remaining 18 (2 per cent) women, the pregnancies were electively terminated with an alternative method. The median induction-abortion interval was 18.0 hours and 15.0 hours in nulliparous and parous women respectively (P less than 0.0001). The number of pessaries required to induce abortion was not influenced by parity. Significantly more parous women bled more than 500 ml. The incidence of pelvic sepsis (0.1 per cent) and cervical tear (0.1 per cent) was low. Twenty six per cent of women had diarrhoea and 23 per cent vomited following administration of prostaglandin. This study confirmed the efficacy of gemeprost for second trimester termination of pregnancy. This method of termination is safe, non-invasive, simple and has a low complication rate.     

Reductive inactivation of digitoxin by Eubacterium lentum cultures

The obligate anaerobe Eubacterium lentum inactivated the cardiac glycoside digitoxin by reducing the double bond in the lactone ring. This conversion was quantitative when the substrate was incubated at a concentration of 10 micrograms/ml. The reduction reaction coincided with the growth phase of the bacterium. The stereochemical configuration at C-20 of the reduction product dihydrodigitoxin was found to be R. Incubation of digitoxigenin and its mono- and bisdigitoxosides individually with E. lentum led to the formation of their respective dihydro derivatives. The configuration at C-20 of these reduced metabolites was also found to be R.     

Influence of dose and route of administration of bovine follicular fluid and the suppressive effect of purified bovine inhibin (Mr 31,000) on plasma FSH concentrations in ovariectomized ewes

Ovariectomized Merino ewes were used to develop an in-vivo bioassay for purified bovine inhibin of Mr 31,000. Various doses (0.25, 0.5, 1 or 2 ml) of bovine follicular fluid, given either by the intravenous (i.v.) or intracarotid route (i.c.) resulted in significant linear dose-related suppression of plasma FSH and interval to maximum suppression. Control ewes (1.0 ml steer plasma) showed no significant change in FSH over the same period. Doses of 470 and 2590 U of pure inhibin given i.v. caused a significant suppression of FSH in plasma in all ewes. The in-vivo potency estimate of the high dose (2760 U, 1420-4690 fiducial limits) agreed well with the in-vitro assay of potency. There were no significant changes observed in mean plasma LH after treatment with the higher dose of pure inhibin. There were no rebound effects of treatment with bovine follicular fluid or pure inhibin on FSH concentrations above that of controls. It is concluded that the form of bovine inhibin of Mr 31,000, which is believed to be the predominant circulating form, is biologically active when administered in vivo.     

Purification and Characterization of Cytosolic NADP Specific Isocitrate Dehydrogenase from Pisum sativum

Cytosolic NADP-specific isocitrate dehydrogenase was isolated from leaves of Pisum sativum. The purified enzyme was obtained by ammonium sulfate fractionation, ion exchange, affinity, and gel filtration chromatography. The purification procedure yields greater than 50% of the total enzyme activity originally present in the crude extract. The enzyme has a native molecular weight of 90 kilodaltons and is resolved into two catalytically active bands by isoelectric focusing. Purified NADP-isocitrate dehydrogenase exhibited K_m values of 23 micromolar for dl-isocitrate and 10 micromolar for NADP, and displayed optimum activity at pH 8.5 with both Mg²⁺ and Mn²⁺.     

Protein Synthesis in Bromegrass (Bromus inermis Leyss) Cultured Cells during the Induction of Frost Tolerance by Abscisic Acid or Low Temperature

Bromus inermis Leyss cell cultures treated with 75 micromolar abscisic acid (ABA) at both 23 and 3°C developed more freezing resistance than cells cultured at 3°C. Protein synthesis in cells induced to become freezing tolerant by ABA and low temperature was monitored by [¹⁴C]leucine incorporation. Protein synthesis continued at 3°C, but net cell growth was stopped. Most of the major proteins detected at 23°C were synthesized at 3°C. However, some proteins were synthesized only at low temperatures, whereas others were inhibited. ABA showed similar effects on protein synthesis at both 23 and 3°C. Comparative electrophoretic analysis of [¹⁴C]leucine labeled protein detected the synthesis of 19, 21 and 47 kilodalton proteins in less than 8 hours after exposure to exogenous ABA. Proteins in the 20 kilodalton range were also synthesized at 3°C. In addition, a 31 kilodalton protein band showed increased expression in freezing resistant ABA treated cultures after 36 hours growth at both 3 and 23°C. Quantitative analysis of [¹⁴C]leucine labeled polypeptides in two-dimensional gels confirmed the increased expression of the 31 kilodalton protein. Two-dimensional analysis also resolved a 72 kilodalton protein enriched in ABA treated cultures and identified three proteins (24.5, 47, and 48 kilodaltons) induced by low temperature growth.