scholarly journals MACROPHAGE-MELANOMA CELL HETEROKARYONS

1971 ◽  
Vol 134 (4) ◽  
pp. 935-946 ◽  
Author(s):  
Saimon Gordon ◽  
Zanvil Cohn
Keyword(s):  
Protein Synthesis ◽  
Dna Synthesis ◽  
Melanoma Cell ◽  
Melanoma Cells ◽  
Irreversible Inhibitor ◽  
Cell Pretreatment ◽  
Lag Period ◽  
Inhibitor Of Protein Synthesis ◽  
Physical Changes ◽  
The Relationship

Dormant macrophage nuclei initiate DNA synthesis 2–3 hr after fusion of macrophages with exponentially growing melanoma cells. Cycloheximide treatment (1–5 µg/ml) of heterokaryons during the preceding lag period inhibits the initiation of macrophage DNA synthesis, in a reversible fashion. Each type of cell was also treated with streptovitacin A, an irreversible inhibitor of protein synthesis. Pretreatment of the melanoma cells (0.5–2 µg/ml), 1 hr before fusion, inhibited the induction of macrophage DNA synthesis in heterokaryons, whereas pretreatment of macrophages (1–20 µg/ml) had no effect. Melanoma cell pretreatment reduced the incorporation of leucine-3H into the cytoplasm and nuclei of heterokaryons, whereas macrophage pretreatment had no effect. These experiments suggested that melanoma proteins played an important role in the initiation of macrophage DNA synthesis. The relationship between the melanoma cell cycle and macrophage DNA synthesis was studied with synchronous melanoma cells. If the melanoma cells were in S phase at the time of fusion, macrophage DNA synthesis occurred 2 hr later. However, the fusion of melanoma cells in G1 delayed macrophage DNA synthesis until the melanoma nuclei had entered S. Experiments with actinomycin and cycloheximide showed that RNA and protein, essential to achieve DNA synthesis in the macrophage nucleus, were made during late G1 as well as S. Melanoma cells and macrophages differ in their radiolabeled acid-soluble products after incubation in thymidine-3H. Thymidine taken up by the macrophage remained unphosphorylated, whereas it was recovered mainly as thymidine triphosphate from melanoma cells. These findings, as well as those reported previously, suggest that the melanoma cell provides the RNA, protein, and precursors which initiate macrophage DNA synthesis. In the absence of a requirement for new macrophage RNA and protein synthesis, other changes must be responsible for the 2 hr delay in DNA synthesis. These may involve physical changes in DNA, associated with swelling, as well as the transport of melanoma products into the macrophage nucleus.

scholarly journals LIMITED DNA SYNTHESIS IN THE ABSENCE OF PROTEIN SYNTHESIS IN PHYSARUM POLYCEPHALUM

1966 ◽  
Vol 31 (3) ◽  
pp. 577-583 ◽  
Author(s):  
J. E. Cummins ◽  
H. P. Rusch
Keyword(s):  
Protein Synthesis ◽  
Dna Replication ◽  
Dna Synthesis ◽  
Physarum Polycephalum ◽  
Nuclear Dna ◽  
S Phase ◽  
Early Prophase ◽  
Late Prophase ◽  
Removal Experiments ◽  
Inhibitor Of Protein Synthesis

Actidione (cycloheximide), an antibiotic inhibitor of protein synthesis, blocked the incorporation of leucine and lysine during the S phase of Physarum polycephalum. Actidione added during the early prophase period in which mitosis is blocked totally inhibited the initiation of DNA synthesis. Actidione treatment in late prophase, which permitted mitosis in the absence of protein synthesis, permitted initiation of a round of DNA replication making up between 20 and 30% of the unreplicated nuclear DNA. Actidione treatment during the S phase permitted a round of replication similar to the effect at the beginning of S. The DNA synthesized in the presence of actidione was replicated semiconservatively and was stable through at least the mitosis following antibiotic removal. Experiments in which fluorodeoxyuridine inhibition was followed by thymidine reversal in the presence of actidione suggest that the early rounds of DNA replication must be completed before later rounds are initiated.

scholarly journals Investigations of the Mode of Action and Resistance Development of Cadazolid, a New Antibiotic for Treatment of Clostridium difficile Infections

2013 ◽  
Vol 58 (2) ◽  
pp. 901-908 ◽  
Author(s):  
Hans H. Locher ◽  
Patrick Caspers ◽  
Thierry Bruyère ◽  
Susanne Schroeder ◽  
Philippe Pfaff ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Clostridium Difficile ◽  
Dna Synthesis ◽  
Mode Of Action ◽  
Cross Resistance ◽  
Resistance Development ◽  
Content Type ◽  
Inhibition Of Protein Synthesis ◽  
Inhibitor Of Protein Synthesis ◽  
Inhibition Of Dna Synthesis

ABSTRACTCadazolid is a new oxazolidinone-type antibiotic currently in clinical development for the treatment ofClostridium difficile-associated diarrhea. Here, we report investigations on the mode of action and the propensity for spontaneous resistance development inC. difficilestrains. Macromolecular labeling experiments indicated that cadazolid acts as a potent inhibitor of protein synthesis, while inhibition of DNA synthesis was also observed, albeit only at substantially higher concentrations of the drug. Strong inhibition of protein synthesis was also obtained in strains resistant to linezolid, in agreement with low MICs against such strains. Inhibition of protein synthesis was confirmed in coupled transcription/translation assays using extracts from differentC. difficilestrains, including strains resistant to linezolid, while inhibitory effects in DNA topoisomerase assays were weak or not detectable under the assay conditions. Spontaneous resistance frequencies of cadazolid were low in all strains tested (generally <10−10at 2× to 4× the MIC), and in multiple-passage experiments (up to 13 passages) MICs did not significantly increase. Furthermore, no cross-resistance was observed, as cadazolid retained potent activity against strains resistant or nonsusceptible to linezolid, fluoroquinolones, and the new antibiotic fidaxomicin. In conclusion, the data presented here indicate that cadazolid acts primarily by inhibition of protein synthesis, with weak inhibition of DNA synthesis as a potential second mode of action, and suggest a low potential for spontaneous resistance development.

scholarly journals Induction of human choriogonadotropin in HeLa-cell cultures by aliphatic monocarboxylates and inhibitors of deoxyribonucleic acid synthesis

1977 ◽  
Vol 166 (2) ◽  
pp. 265-274 ◽  
Author(s):  
Nimai K. Ghosh ◽  
Adriana Rukenstein ◽  
Rody P. Cox
Keyword(s):  
Protein Synthesis ◽  
Hela Cell ◽  
Dna Synthesis ◽  
Hela Cells ◽  
Fold Increase ◽  
Short Chain Fatty Acids ◽  
Acid Synthesis ◽  
Hydroxyl Groups ◽  
Β Subunit ◽  
Inhibitor Of Protein Synthesis

The ectopic production of the glycopeptide hormone human placental choriogonadotropin by HeLa65 cells was measured by radioimmunoassay with antiserum against the β-subunit of choriogonadotropin and with the125I-labelled β-subunit as a tracer antigen. Choriogonadotropin synthesis was markedly (500-fold) stimulated by sodium butyrate. Kinetic studies and the use of an inhibitor of protein synthesis, cycloheximide, indicated that protein synthesis was required for this induction. Investigation of the efficiency of 22 aliphatic short-chain fatty acids and derivatives in causing increased choriogonadotropin synthesis by HeLa cells showed stringent structural requirements. Induction of choriogonadotropin synthesis in HeLa cells was not restricted to butyrate. Other aliphatic acids (propionate, isobutyrate, valerate and hexanoate) were also capable of inducing choriogonadotropin synthesis at 10–50% of the efficiency of butyrate. Hydroxy derivatives of monocarboxylate inducers, related mono- and di-carboxylic acids, alcohols, amines, ketones, esters and sulphoxide were ineffective in increasing choriogonadotropin production by HeLa cells. A saturated C4 straight-chain acid without substituent hydroxyl groups but with a methyl group at one end and a carboxyl moiety at the other appeared to be most efficient in activating choriogonadotropin production. A second clonal line of HeLa cells, HeLa71, showed a higher constitutive synthesis of choriogonadotropin than HeLa65 cells, which was also markedly increased by butyrate. Butyrate and other aliphatic monocarboxylate inducers of choriogonadotropin synthesis inhibited HeLa-cell growth and DNA synthesis. This inhibition of DNA replication may be related to the mechanism of choriogonadotropin synthesis, since two well-characterized anti-neoplastic inhibitors of DNA synthesis, hydroxyurea and 1-β-d-arabinofuranosylcytosine, also stimulated a 300-fold increase in choriogonadotropin synthesis in HeLa cells and were synergistic with butyrate in promoting choriogonadotropin synthesis. Thus activation in tumour cells of genes normally expressed by foetal tissue and the consequent ectopic synthesis of polypeptide hormones may require neither cell division nor DNA synthesis.

Expression of tetracycline resistance mediated by an R-factor in Escherichia coli 15T−

1974 ◽  
Vol 20 (4) ◽  
pp. 545-549
Author(s):  
M. L. Braun ◽  
D. W. Ferrell ◽  
J. L. Ott
Keyword(s):  
Escherichia Coli ◽  
Protein Synthesis ◽  
Dna Synthesis ◽  
Tetracycline Resistance ◽  
Cell Multiplication ◽  
Cell Numbers ◽  
R Factor ◽  
Lag Period ◽  
Low Levels ◽  
H Protein

Streptomycin and tetracycline resistance were carried on an R-factor in Escherichia coli 15T−R64-11 and 15T−NArR64-11. When growth of these strains was studied in the presence of the antibiotic, tetracycline caused a long lag period before growth occurred. During this lag, cell numbers decreased before cells multiplied. The lag period was eliminated by pretreatment of the inoculum with low levels of the antibiotic. Macromolecular synthesis was measured by incorporation of radioactive precursors into TCA-precipitable material in the absence and presence of tetracycline. In the absence of the antibiotic, RNA and protein synthesis were immediate while DNA synthesis occurred after a 15-min lag. During the lag period caused by tetracycline RNA synthesis was initiated but stopped after 1 h, and then resumed at 4 h. Protein synthesis was evident at 2 h while DNA synthesis occurred simultaneously with cell multiplication. Tetracycline also caused a long lag period when lactose was used as the energy source, but cell numbers did not decrease during this lag.

scholarly journals IRF4-activated TEX41 promotes the malignant behaviors of melanoma cells by targeting miR-103a-3p/C1QB axis

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Yingna Zheng ◽  
Wu Zhou ◽  
Min Li ◽  
Ruixue Xu ◽  
Shuai Zhang ◽  
...  
Keyword(s):  
Melanoma Cell ◽  
Melanoma Cells ◽  
Migration And Invasion ◽  
Transcriptional Level ◽  
Potential Therapeutic Target ◽  
Expression Of Genes ◽  
Melanoma Cell Proliferation ◽  
Non Coding Rnas ◽  
The Relationship

Abstract Background Malignant melanoma is an aggressive skin cancer and a tumor of melanocytic origin. Recent studies have suggested that long non-coding RNAs (lncRNAs) play crucial regulatory roles in multiple malignancies, including melanoma. Testis expressed 41 (TEX41) is a relatively new lncRNA whose mechanism in melanoma remains vague. Aims This study aimed to explore the role and specific mechanism of TEX41 in melanoma. Methods The expression of genes involved in this study was determined by qRT-PCR. Functional assays were conducted to analyze the role of relevant genes in melanoma cells. The interaction between TEX41 promoter and IRF4 as well as the relationship among TEX41, miR-103a-3p and C1QB was verified by mechanism assays. Results IRF4 up-regulated TEX41 at the transcriptional level in melanoma cells. TEX41 knockdown hindered melanoma cell proliferation, migration and invasion while promoting cell apoptosis. TEX41 bound to miR-103a-3p and regulated C1QB. The suppressive impact of TEX41 depletion on melanoma cell malignant behaviors could be counteracted by miR-103a-3p inhibition or C1QB overexpression. Moreover, IRF4 could facilitate melanoma cell growth via up-regulating C1QB. Conclusions IRF4-activated TEX41 sequestered miR-103a-3p and modulated C1QB to promote melanoma cell malignant behaviors, for which TEX41 might be regarded as a potential therapeutic target for melanoma.

scholarly journals Multifunctional bioscaffolds for 3D culture of melanoma cells reveal increased MMP activity and migration with BRAF kinase inhibition

2015 ◽  
Vol 112 (17) ◽  
pp. 5366-5371 ◽  
Author(s):  
Jennifer L. Leight ◽  
Emi Y. Tokuda ◽  
Caitlin E. Jones ◽  
Austin J. Lin ◽  
Kristi S. Anseth
Keyword(s):  
Drug Treatment ◽  
Metastatic Melanoma ◽  
Proteolytic Activity ◽  
Melanoma Cell ◽  
3D Culture ◽  
Melanoma Cells ◽  
Functional Change ◽  
Three Dimensions ◽  
Preclinical Research ◽  
The Relationship

Matrix metalloproteinases (MMPs) are important for many different types of cancer-related processes, including metastasis. Understanding the functional impact of changes in MMP activity during cancer treatment is an important facet not typically evaluated as part of preclinical research. With MMP activity being a critical component of the metastatic cascade, we designed a 3D hydrogel system to probe whether pharmacological inhibition affected human melanoma cell proteolytic activity; metastatic melanoma is a highly aggressive and drug-resistant form of skin cancer. The relationship between MMP activity and drug treatment is unknown, and therefore we used an in situ fluorogenic MMP sensor peptide to determine how drug treatment affects melanoma cell MMP activity in three dimensions. We encapsulated melanoma cells from varying stages of progression within PEG-based hydrogels to examine the relationship between drug treatment and MMP activity. From these results, a metastatic melanoma cell line (A375) and two inhibitors that inhibit RAF (PLX4032 and sorafenib) were studied further to determine whether changes in MMP activity led to a functional change in cell behavior. A375 cells exhibited increased MMP activity despite an overall decrease in metabolic activity with PLX4032 treatment. The changes in proteolytic activity correlated with increased cell elongation and increased single-cell migration. In contrast, sorafenib did not alter MMP activity or cell motility, showing that the changes induced by PLX4032 were not a universal response to small-molecule inhibition. Therefore, we argue the importance of studying MMP activity with drug treatment and its possible implications for unwanted side effects.

scholarly journals Suppression of the formation of polyamines and macromolecules by dl-α-difluoromethylornithine and methylglyoxal bis(guanylhydrazone) in phytohaemagglutinin-activated human lymphocytes

1979 ◽  
Vol 178 (1) ◽  
pp. 109-117 ◽  
Author(s):  
Juhani Jänne ◽  
Tapani Hovi ◽  
Erkki Hölttä
Keyword(s):  
Protein Synthesis ◽  
Dna Synthesis ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Peripheral Blood Lymphocytes ◽  
Irreversible Inhibitor ◽  
Polyamine Biosynthesis ◽  
Inhibition Of Protein Synthesis ◽  
Dose Dependent Decrease

1. The activation of human peripheral blood lymphocytes by phytohaemagglutinin in vitro was accompanied by striking increases in the concentrations of the natural polyamines putrescine, spermidine and spermine. 2. The enhanced accumulation of polyamines could be almost totally abolished by dl-α-difluoromethylornithine, a newly discovered irreversible inhibitor of l-ornithine decarboxylase (EC 4.1.1.17), or by methylglyoxal bis(guanylhydrazone) {1,1′-[(methylethanediylidene)dinitrilo]diguanidine}, an inhibitor of S-adenosyl-l-methionine decarboxylase (EC 4.1.1.50). The inhibition of polyamine accumulation was associated with a marked suppression of DNA synthesis, which was partially or totally reversed by low concentrations of exogenous putrescine, spermidine, spermine and cadaverine and by higher concentrations of 1,3-diaminopropane. 3. In contrast with some earlier studies, we found that methylglyoxal bis(guanylhydrazone), at concentrations that were sufficient to prevent polyamine accumulation, also caused a clear inhibition of protein synthesis in the activated lymphocytes. Similar results were obtained with difluoromethylornithine. The decrease in protein synthesis caused by both compounds preceded the impairment of DNA synthesis. The inhibition of protein synthesis by difluoromethylornithine was fully reversed by exogenous putrescine, spermidine and spermine, and that caused by methylglyoxal bis(guanylhydrazone) by spermidine and spermine. In further support of the idea that the inhibition of protein synthesis by these compounds was related to the polyamine depletion, we found that difluoromethylornithine caused a dose-dependent decrease in the incorporation of [14C]leucine into lymphocyte proteins which closely correlated with the decreased concentrations of cellular spermidine. 4. Difluoromethylornithine and methylglyoxal bis(guanylhydrazone) also elicited a variable depression in the incorporation of [3H]uridine and [14C]adenine into total RNA. The apparent turnover of lymphocyte RNA remained essentially unchanged in spite of severe polyamine depletion brought about by difluoromethylornithine. 5. The present results, as well as confirming the anti-proliferative action of the inhibitors of polyamine biosynthesis, suggest that polyamine depletion may interfere with reactions at different levels of gene expression.

scholarly journals STUDIES ON THE MECHANISM OF ACTION OF PEDERINE

1968 ◽  
Vol 36 (3) ◽  
pp. 485-496 ◽  
Author(s):  
Agnese Brega ◽  
Arturo Falaschi ◽  
Luigi De Carli ◽  
Mario Pavan
Keyword(s):  
Protein Synthesis ◽  
Dna Synthesis ◽  
Cytotoxic Effect ◽  
Rna Synthesis ◽  
Cultured Cells ◽  
Specific Inhibitor ◽  
Cytological Examination ◽  
Effect Analysis ◽  
Inhibitor Of Protein Synthesis

Pederine, a drug extracted from the coleopter Paederus fuscipes, inhibits the growth of in vitro cultured cell lines at concentrations of the order of 1.5 nanogram/ml. Cytological examination shows a generalized cytotoxic effect. Analysis of macromolecular syntheses by the use of radioactive precursors shows that pederine causes an almost immediate block of protein and DNA synthesis, without affecting RNA synthesis. The effects on the synthesis of the two types of macromolecules remain nearly simultaneous even at the lowest active concentrations of pederine. Studies with cell-free systems show that the drug inhibits protein synthesis, whereas it is ineffective on the DNA-polymerizing activity. It seems, therefore, that the drug acts primarily on the amino acid-polymerizing system, and that the effect on DNA is secondary. This idea is strengthened by the observation that puromycin, a specific inhibitor of protein synthesis, also affects promptly DNA synthesis of in vitro cultured cells. Other authors have shown the same phenomenon with a number of inhibitors of protein synthesis; the properties of pederine support, therefore, the view that continuous protein synthesis is necessary for the maintenance of DNA replication in higher organisms.

Proline Accumulation in Tubers of Jerusalem Artichoke

1977 ◽  
Vol 4 (1) ◽  
pp. 51 ◽  
Author(s):  
L Wright ◽  
P Wrench ◽  
RW Hinde ◽  
CJ Brady
Keyword(s):  
Jerusalem Artichoke ◽  
Competitive Inhibitor ◽  
Proline Accumulation ◽  
Free Proline ◽  
Higher Plant ◽  
Tuber Tissue ◽  
Arginine Catabolism ◽  
Lag Period ◽  
Inhibitor Of Protein Synthesis ◽  
The Relationship

When separated whole tubers of Helianthus tuberosus L. were dried, and when tuber slices were plasmolysed in mannitol solutions, free proline accumulated in the tuber tissue. The amount of proline accumulated in tuber slices increased with the concentration of mannitol in the plasmolysing solution. A lag of approximately 24 h preceded proline accumulation in plasmolysed slices. During the lag period, the content of free arginine, a major component of the free amino acid pool, decreased by at least 50%. L-Indospicine, a competitive inhibitor of arginase (L-arginine amidinohydrolase EC 3.5.3.1) inhibited arginine catabolism in slices and also inhibited proline accumulation. On the other hand, cycloheximide, an inhibitor of protein synthesis, did not inhibit arginine loss or proline accumulation. An increase in the concentration of mannitol required for plasmolysis accompanied an increase in free proline in whole, dormant tubers stored at 4°C. These tubers are an example of a non-photosynthetic higher plant tissue in which free proline accumulates when the water potential decreases. The relationship between arginine catabolism and proline accumulation is discussed.

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