Expression of antibody cDNA in murine myeloma cells: possible involvement of additional regulatory elements in transcription of immunoglobulin genes

Expression vectors for cDNA of the κ and λ1 chains of a monoclonal antibody directed against creatine kinase were introduced into murine myeloma cells. κ and γ1 cDNA were either under the control of the SV40 early promoter or of the cognate promoters and enhancers of the light- and heavy-chain genes. Secretion of immuno-reactive κ and γ1 chains into the culture medium was demonstrated with the SV40 promoter as well as with the cognate promoters. Expression of y 1 cDNA with the SV40 early promoter was about twice as high as with the heavy-chain promoter and enhancer. Expression of κ cDNA under the control of the S V40 early promoter was about 17 times higher than with the light-chain promoter and enhancer. These expression levels were compared to those of a genomic immunoglobulin (Ig) κ determinant, including introns. Such an entire κ gene led to expression of the light chain at levels double those with the κ cDNA construction using the SV40 promoter and about 35 times as high when using κ cDNA and the cognate promoter and enhancer. This result might indicate that, besides the cognate promoter and enhancer elements, other intragenic elements are involved in the regulation of Ig expression. However, the SV40 early promoter seems to be able to compensate for the absence of these postulated regulatory elements probably located in the introns.     

Influence of α-interferon therapy on blood lymphoid cells

Summary The influence of natural -interferon (-IFN) therapy (3×10⁶ units i.m. daily) on blood lymphoid cells was studied in 20 patients with gynecological neoplasias (7 patients with condylomata accuminata and 13 patients with ovarian carcinoma). There was a statistically significant increase in the intracellular levels of 2'–5'oligoadenylate synthese 1 day after the first injection of IFN and with few exceptions this activity remained increased during 3 months of treatment. In most of the patients, the capacity of blood lymphoid cells to produce IgA, IgG, and IgM following stimulation with pokeweed mitogen was decreased 1 day after the first injection of IFN and with few exceptions it remained low during 6 months of IFN therapy. In most patients there was a decrease in the capacity of lymphoid cells to act as stimulator or responder cells in a mixed lymphocyte culture during IFN therapy. The -IFN therapy had no major influence on the response of lymphoid cells to mitogens. We conclude, that neither this nor our previous studies on the influence of IFN therapy on immunological functions have given support to the hypothesis that the antitumor action of IFN is mediated by the immune system.     

Immunohistochemical localization of tissue-type plasminogen activator in ovaries before and after induced and spontaneous ovulation in the rat

Summary The observation that tissue-type plasminogen activator (tPA) activity increased dramatically in preovulatory follicles has led to the hypothesis that plasminogen activation is causally related to follicle rupture. With immunohistochemistry, we have studied the appearance of tPA in ovaries of immature rats induced to ovulate and in adult cycling rats. Treatment of immature female rats with a single dose of pregnant mare serum gonadotropin (PMSG) induced follicular maturation. A subsequent human chorionic gonadotropin (hCG) injection resulted in follicle rupture 12–14 h later. PMSG treatment alone did not induce appearance of tPA-immunoreactive cells in any ovarian compartment. After hCG stimulation, however, theca cells, granulosa cells, and oocytes of pre- and postovulatory follicles displayed distinct tPA immunoreactivity. Fibroblastlike cells in the theca layers and tunica albuginea of the follicle apex also demonstrated localized cytoplasmic tPA reactivity. In addition to tPA synthesis in preovulatory follicles, hCG also induced tPA staining in the theca (but not granulosa) layers of non-ovulatory follicles. At 24 h after hCG treatment, there was a marked tPA staining in developing corpora lutea, ovulated ova, and oviductal epithelium. Ovaries from regularly cycling adult rats displayed a similar ovulation-related pattern of tPA immunostaining. The appearance of tPA in different cell types of the preovulatory follicle and in the fibroblast-like cells at the follicle apex, strengthens the hypothesis of a direct involvement of tPA in follicle rupture. Presence of tPA in postovulatory oocytes, cumulus cells, and surrounding oviductal epithelium may also indicate a role for tPA in the transfer of eggs in the oviduct.This work was supported by NIH Research Grants HD-14084; 12303     

Differentiation of embryonic chick sympathetic neurons in vivo: ultrastructure,and quantitative determinations of catecholamines and somatostatin

Summary The ultrastructural and transmitter development of lumbar sympathetic ganglia was studied in embryonic day-6 through-18 chick embryos. At embryonic day 6, ganglia are populated by two morphologically distinct types of neuronal cells and Schwann cell precursors. The neuronal populations basically comprise a granule-containing cell and a developing principal neuron. Granule-containing cells have, an irregularly shaped or oval nucleus with small clumps of chromatin attached to the inner nuclear membrane and numerous large (up to 300 nm) membrane-limited granules. Developing principal neurons display a more rounded vesicular nucleus with evenly distributed chromatin, prominent nucleoli, more developed areas of Golgi complexes, and rough endoplasmic reticulum and large dense-core vesicles up to 120 nm in diameter. There are granule-containing cells with fewer and smaller granules which still display the nucleus typical for granule-containing cells. These granule-containing cells may develop toward developing principal neurons or the resting state of granule-containing cells found in older ganglia. Both granule-containing cells and developing principal neurons proliferate and can undergo degeneration. At embryonic day 9 there are far more developing principal neurons than granule-containing cells. Most granule-containing cells have very few granules. Mitotic figures and signs of cell degeneration are still apparent. Synapse-like terminals are found on both developing principal neurons and granule-containing cells. Ganglionic development from embryonic day 11 through 18 comprises extensive maturation of developing principal neurons and a numerical decline of granule-containing cells. Some granule-containing cells with very few and small granules still persist at embryonic day 18. The mean catecholamine content per neuron increases from 0.044 femtomol at embryonic day 7 to 0.22 femtomol at embryonic day 15. Concomitantly, there is a more than 6-fold increase in tyrosine hydroxylase activity. Adrenaline has a 14% share in total catecholamines at embryonic day 15. Somatostatin levels are relatively high at embryonic day 7 (1.82 attomol per neuron) and are 10-fold reduced by embryonic day 15. Our results suggest the presence of two morphologically distinct sympathetic neuronal precursors at embryonic day 6: one with a binary choice to become a principal neuron or to die, the other one, a granule-containing cell, which alternatively may develop into a principal neuron, acquire a resting state or die.     

Dopamine Receptor Stimulation Does Not Affect Phosphoinositide Hydrolysis in Slices of Rat Striatum

The effect of dopamine receptor stimulation on the accumulation of labelled inositol phosphates in rat striatal slices under basal and stimulated conditions was examined following preincubation with [3H]inositol. Incubation of striatal slices with the selective D-1 agonist SKF 38393 or the selective D-2 agonist LY 171555 for 5 or 30 min did not affect the basal accumulation of labelled inositol mono-, bis-, tris-, and tetrakisphosphate. Resolution by HPLC of inositol trisphosphate into inositol-1,3,4-tris-phosphate and inositol-1,4,5-trisphosphate isomers revealed that under basal conditions dopamine did not influence the accumulation of inositol-1,4,5-trisphosphate. Depolarisation evoked by KCl, or addition of the muscarinic receptor agonist carbachol, produced a marked increase in the accumulation of labelled inositol phosphates in both the presence and absence of lithium. Addition of dopamine did not reduce the ability of KCl or carbachol to increase inositol phospholipid hydrolysis. In the presence of lithium, dopamine (100 microM) enhanced KCl-stimulated inositol phospholipid hydrolysis, but this effect appears to be mediated by alpha 1 adrenoceptors because it was blocked by prazosin. SKF 38393 (10 microM) or LY 171555 (10 microM) also did not affect carbachol-stimulated inositol phospholipid hydrolysis. These data, in contrast to recent reports, suggest that striatal dopamine receptors do not appear to be linked to inositol phospholipid hydrolysis.     

A study on the mechanism of the vanadate-dependent NADH oxidation

The mechanism of the vanadate (V(_v))-dependent oxidation of NADH was different in phosphate buffers and in phosphate-free media. In phosphate-free media (aqueous medium or HEPES buffer) the vanadyl (V(_v)) generated by the direct V(_v)-dependent oxidation of NADH formed a complex with V(_v). In phosphate buffers V(_v) autoxidized instead of forming a complex with V(_v). The generated superoxide radical (O₂⁻) initiated, in turn, a high-rate free radical chain oxidation of NADH. Phosphate did not stimulate the V(_v)-dependent NADH oxidation catalyzed by O₂⁻-generating systems. Monovanadate proved to be a stronger catalyzer of NADH oxidation as compared to polyvanadate.     

Nerve cell and nematocyte production in Hydra is deregulated by lithium ions

Summary LiCl, a well-known vegetalising agent, interferes with the commitment of stem cells to nerve cells and nematocytes in Hydra attenuata. Treatment with 20 mM LiCl inhibits commitment to nerve cells, treatment with 1 mM LiCl inhibits commitment to nematocytes. However, LiCl does not prevent stem cells committed to the nematocyte pathway from dividing and differentiating into nests of nematocytes. Following LiCl treatment, determination to nerve cells and nematocytes is triggered again. Commitment to nerve cells is strongly stimulated within the first 3 h following pulse treatment with LiCl if the animals have been fed immediately prior to treatment. In Hydra exposed to LiCl for 10 days the stem cell density is reduced by at least 90% of the initial value, and nematocytes are almost completely missing, whereas the density of nerve cells is within the normal range in animals with normal morphology. Animals which developed a transverse constriction in the middle of the body axis contain a 1.7-fold higher nerve cell density in the lower part than is observed in control animals.     

Studies on the effect of NAD(H) on nitrogenase activity in Rhodospirillum rubrum

The effect of NAD(P) and analogs of this nucleotide on nitrogenase activity in Rhodospirillum rubrum has been studied. Addition of NAD⁺ to nitrogen fixing Rsp. rubrum leads to inhibition of nitrogenase. NADP⁺ has the same effect but NADH or analogs modified in the nicotinamide portion do not cause inhibition. In contrast to ammonium ions, addition of NAD⁺ leads to inhibition of nitrogenase in cells that have been N-starved under argon. The inhibitory effect of NAD⁺ is more pronounced at lower light intensities. Addition of NAD⁺ also leads to inhibition of glutamine synthetase, a phenomenon also occurring when “switchoff” is produced by the addition of effectors such as ammonium ions or glutamine. It is also shown that NAD⁺ is taken up by Rsp. rubrum cells.     

A nomenclature for the genes encoding the chlorophylla/b-binding proteins of higher plants

We propose a nomenclature for the genes encoding the chlorophylla/b-binding proteins of the light-harvesting complexes of photosystem I and II. The genes encoding LHC I and LHC II polypeptides are namedLhca1 throughLhca4 andLhcb1 throughLhcb6, respectively. The proposal follows the general format recommended by the Commision on Plant Gene Nomenclature. We also present a table for the conversion of old gene names to the new nomenclature.