Ribosomal protein S12 as a site for streptomycin resistance in Nicotiana chloroplasts

Summary A streptomycin resistant Nicotiana plastome mutant, X/str ^R6, was subjected to molecular analysis. In this mutant, a single nucleotide transition, C » T, in the chloroplast gene for ribosomal protein S12 alters codon 90 from proline to serine while the nucleotide sequence of the chloroplast 16 S rRNA gene is identical to that of the wild type. Mutant X/str ^R6 thus differs from several previously reported streptomycin resistant chloroplast mutants which are altered in the gene for 16 S rRNA.     

Weather- and population density-induced infantilism in the landsnailTheba pisana in a semi-arid climate

Natural populations of the landsnailTheba pisana (Pulmonata: Helicidae) were studied in the Mediterranean coastal plains of Israel. The life cycle is annual. Egg-laying occurs in the winter and the descendants grow fast during the spring, except for a part of the population the ceases growing. These individuals, termed infantiles, retain immature size and shape and a rudimentary status of the genital system. The percentage of infantilism in the population is positively related to the density of the snail population in the winter, and is negatively related to the humidity of weather in the spring. A natural control mechanism ofT. pisana populations is proposed: (a) in a dense population of young snails infantilism prevents most of them from becoming parents and an over-sized population the following year; (b) in a humid spring a fall in the rate of infantilism enlarges the population size, thus compensating for reduced egg-laying in winter.     

Calcium mobilizing hormones activate the plasma membrane Ca2+ pump of pancreatic acinar cells

Summary ⁴⁵Ca fluxes and free-cytosolic Ca²⁺ ([Ca²⁺] _i ) measurements were used to study the effect of Ca²⁺-mobilizing hormones on plasma membrane Ca²⁺ permeability and the plasma membrane Ca²⁺ pump of pancreatic acinar cells. We showed before (Pandol, S.J., et al., 1987.J. Biol. Chem. 262:16963–16968) that hormone stimulation of pancreatic acinar cells activated a plasma membrane Ca²⁺ entry pathway, which remains activated for as long as the intracellular stores are not loaded with Ca²⁺. In the present study, we show that activation of this pathway increases the plasma membrane Ca²⁺ permeability by approximately sevenfold. Despite that, the cells reduce [Ca²⁺]i back to near resting levels. To compensate for the increased plasma membrane Ca²⁺ permeability, a plasma membrane Ca²⁺ efflux mechanism is also activated by the hormones. This mechanism is likely to be the plasma membrane Ca²⁺ pump. Activation of the plasma membrane Ca²⁺ pump by the hormones is time dependent and 1.5–2 min of cell stimulation are required for maximal Ca²⁺ pump activation. From the effect of protein kinase inhibitors on hormone-mediated activation of the pump and the effect of the phorbol ester 12-0-tetradecanoyl phorbol, 13-acetate (TPA) on plasma membrane Ca⁺ efflux, it is suggested that stimulation of protein kinase C is required for the hormone-dependent activation of the plasma membrane Ca²⁺ pump.     

Chemo-adoptive immunotherapy of nude mice implanted with human colorectal carcinoma and melanoma cell lines

Summary The antitumor effects of chemotherapy, recombinant human interleukin-2 (IL-2), recombinant human interferon A/D (IFN), allogeneic human lymphokine-activated killer (LAK) cells, and antitumor monoclonal antibody (mAb), administered alone and in various combinations, were tested in athymic nude mice carrying human tumor xenografts. Treatment began 6–18 days after i.v. or i.p. inoculation of colorectal carcinoma or melanoma cell lines, when macroscopic growths were evident. Chemotherapy consisted of two or three courses of 5-fluorouracil (5-FU) or dacarbazine. IL-2 and/or IFN were administered three to five times weekly for 1–3 weeks, usually starting 2–5 days after chemotherapy. Human LAK cells were infused once or twice weekly for 2 or 3 weeks concurrently with IL-2. In some experiments, murine anticolorectal carcinoma mAb (SF25) was administered. In both tumor systems, chemotherapy alone or immunotherapy alone (IL-2, IL-2 + LAK cells, IFN, IL-2 + IFN ± LAK cells) had little or no therapeutic effects. Additive effects were obtained by combining chemotherapy with IL-2 and LAK cells or with IL-2 and IFN. In the majority of the experiments, the most effective combination was chemotherapy + IL-2 + IFN + LAK cells. Treatment with mAb was beneficial in the colorectal carcinoma system when combined with 5-FU + IL-2 or 5-FU + IL-2 + IFN. Homing experiments with radiolabeled human and mouse LAK cells injected i.v. showed increased early accumulation in the liver and lungs, whereas freshly explanted mouse splenocytes localized mostly in the spleen and liver. The tissue distribution pattern of human LAK cells was similar in normal and tumor-bearing mice (with lung metastases). These findings suggest that combination of chemotherapy with cytokines and LAK cells can be partially effective for advanced solid human tumors even in the absence of the host's T-cell immune response. Preliminary experiments showed that tumor-specific, anti-melanoma T-cell clones were effective in local (s.c.) tumor growth inhibition (Winn assay) following coinjection with the autologous tumor cells.     

Immunochemical and biological analysis of pheromone biosynthesis activating neuropeptide in Heliothis peltigera.

This study describes the preparation and characterization of a highly specific antiserum to Helicoverpa zea pheromone biosynthesis activating neuropeptide (Hez-PBAN), and the use of this antiserum, in an enzyme linked immunosorbent assay (ELISA), to determine: a) the content of endogenous PBAN in head extracts of male and female Heliothis peltigera; b) the level of PBAN at different developmental stages; and c) the content of PBAN in four different moth species. Cross-reactivity studies revealed that the antiserum is directed mainly toward the N-terminal region of the neuropeptide, and that it exhibits similar binding affinities toward the oxidized and reduced forms of PBAN. Analysis of PBAN content in head extracts of male and female H. peltigera, at scotophase, revealed the presence of 4.97 and 4.58 pmol, respectively, in 3-day-old moths, and 5.33 and 4.78 pmol, respectively, in 7-day-old moths. The similarity in the content of PBAN at both ages and sexes was in accordance with the amount of pheromonotropic activity in these extracts which stimulated pheromone biosynthesis to a similar level. Analysis of PBAN-like immunoreactivity (IR) in head extracts of H. peltigera larvae and pupae demonstrated the existence of the neuropeptide in the 4th larval instar and continued to increase as a function of development. No IR could be detected in the first three larval instars. The larval and pupal extracts also exerted pheromonotropic activity which followed a similar pattern. The activity in these extracts, however, was considerably lower than that found in adult male and female heads. IR was also detected in head extracts of three other Noctuidae moths: Helicoverpa armigera, Cornutiplusia circumflexa and Spodoptera littoralis, indicating a high degree of chemical and structural similarity of PBAN in these moths.     

Hormone-responsive cells derived from human dental papilla: Characterization in vitro and in vivo in diffusion chambers

Summary Cells of the dental papilla are capable of odontoblastic, fibroblastic, and endothelial differentiation and formation of dentin and the dental pulp. In the present study dental papilla cells, obtained from human tooth buds (HDP cells), were cultured in vitro through 3 to 7 passages. After exposure to prostaglandin E₂ there was a marked decrease in intracellular cyclic AMP (cAMP) levels as compared to hormone-free controls. Parathyroid hormone and calcitonin had stimulatory effects with 1 and 2 log increases in cAMP, respectively. The HDP cells showed moderate activity of alkaline phosphatase, 1 log higher than that of hamster kidney fibroblasts (BHK 13) and 1 log lower than that of osteoblastic osteosarcoma cells (ROS 17/2). When cultured for 4 or 8 wk in diffusion chambers (DC) implanted in athymic mice, many of the HDP cells underwent odontoblastic morphodifferentiation with very long, single processes extending into the matrix. This matrix contained banded and unbanded collagen fibers. Neither light nor electron microscopy of the DC content revealed mineral deposits. These results suggest that HDP cells have an intrinsic potential for partial odontoblastic differentiation; inductive signals like those originating from odontogenic epithelium are probably essential for the completion of hard tissue formation.     

Adaptation and growth of tomato cells on the herbicide 2,6-dichlorobenzonitrile leads to production of unique cell walls virtually lacking a cellulose-xyloglucan network

Suspension-cultured cells of tomato (Lycopersicon esculentum VF 36) have been adapted to growth on high concentrations of 2,6-dichlorobenzonitrile, an herbicide which inhibits cellulose biosynthesis. The mechanism of adaptation appears to rest largely on the ability of these cells to divide and expand in the virtual absence of a cellulose-xyloglucan network. Walls of adapted cells growing on 2,6-dichlorobenzonitrile also differ from nonadapted cells by having reduced levels of hydroxyproline in protein, both in bound and salt-elutable form, and in having a much higher proportion of homogalacturonan and rhamnogalacturonan-like polymers. Most of these latter polymers are apparently cross-linked in the wall via phenolic-ester and/or phenolic ether linkages, and these polymers appear to represent the major load-bearing network in these unusual cell walls. The surprising finding that plant cells can survive in the virtual absence of a major load-bearing network in their primary cell walls indicates that plants possess remarkable flexibility for tolerating changes in wall composition.     

Membrane interactions of the sodium channel S4 segment and its fluorescently-labeled analogues.

A 24-amino acid peptide corresponding to the S4 segment of the sodium channel was synthesized. In order to perform fluorescence energy transfer measurements and to monitor the interaction of the peptide with lipid vesicles, the peptide was selectively labeled with fluorescence probes at either its N- or C-terminal amino acids. The fluorescent emission spectra of 7-nitrobenz-2-oxa-1,3-diazol-4- yl-(NBD-)labeled analogues displayed blue shifts upon binding to small unilamellar vesicles (SUV), reflecting the relocation of the fluorescent probe to an environment of increased apolarity. The results revealed that both the N- and C-terminus of the S4 segment are located within the lipid bilayer. Titration of solutions containing NBD-labeled peptides with SUV was used to generate binding isotherms, from which surface partition constants, in the range of 10(4) M-1, were derived. The shape of the binding isotherms as well as fluorescence energy transfer measurements suggest that aggregation of peptide monomers within the membrane readily occurs in acidic but not in zwitterionic vesicles. Furthermore, the results provide good correlation between the incidence of aggregation in PC/PS vesicles and the ability of the peptides to permeate the vesicle's membrane. However, a transmembrane diffusion potential had no detectable effect on the location of the peptide within the lipid bilayer or on its aggregation state.(ABSTRACT TRUNCATED AT 250 WORDS)