Anticancer Drug Action on Poly(A) Polymerase Activity and Isoforms during Hela and Wish Cell Apoptosis

2000 ◽  
Vol 15 (2) ◽  
pp. 171-178 ◽  
Author(s):  
N.A.A. Balatsos ◽  
M. Havredaki ◽  
C.M. Tsiapalis
Keyword(s):  
Anticancer Agents ◽  
Enzyme Assay ◽  
Polymerase Activity ◽  
Activity Levels ◽  
Immunoblotting Analysis ◽  
A Cell ◽  
Lower Mobility ◽  
Cell Commitment ◽  
Diploid Cells ◽  
Mrna Polyadenylation

Poly(A) polymerase (PAP; EC 2.7.7.19) catalyzes mRNA polyadenylation. Its activity and isoform levels vary during cell cycle transformation and apoptosis. It has become widely accepted that cell death after DNA damage by anticancer agents is primarily the result of apoptosis and that cells able to evade apoptosis will be resistant to cell killing. The therapeutic agents interferon (IFN), 5-fluorouracil (5-FU) and tamoxifen (Tam) with different mechanisms of action mediate both partial dephosphorylation and inactivation of PAP, detected by immunoblotting analysis and PAP enzyme assay, respectively. We examined the apoptotic tendencies of HeLa and WISH cell lines caused by one of the drugs used, 5-FU. The trend in the cells examined, observed by DAPI and/or DNA fragmentation assay, was found to be accompanied by and reversibly related to PAP activity levels and PAP lower mobility phosphorylated forms of 106 and 100 kDa isoforms. Moreover, a cell type-modulated, differential response of HeLa (chemosensitive cells) versus WISH (drug-resistant diploid cells) has been revealed. This finding yields information on the possible use of PAP as a tumor marker involved in cell commitment and/or induction of apoptosis and may help to improve our understanding of tumor cell sensitivity to anticancer agents.

Mating type control in Saccharomyces cerevisiae: a frameshift mutation at the common DNA sequence, X, of the HML alpha locus

1984 ◽  
Vol 4 (1) ◽  
pp. 203-211
Author(s):  
K Tanaka ◽  
T Oshima ◽  
H Araki ◽  
S Harashima ◽  
Y Oshima
Keyword(s):  
Mating Type ◽  
Base Pair ◽  
Reading Frame ◽  
Base Pair Deletion ◽  
A Cell ◽  
Alpha Gene ◽  
The Common ◽  
Type Switch ◽  
Diploid Cells ◽  
Mat Locus

A mutation defective in the homothallic switching of mating type alleles, designated hml alpha-2, has previously been characterized. The mutation occurred in a cell having the HO MATa HML alpha HMRa genotype, and the mutant culture consisted of ca. 10% a mating type cells, 90% nonmater cells of haploid cell size, and 0.1% sporogenous diploid cells. Genetic analyses revealed that nonmater haploid cells have a defect in the alpha 2 cistron at the MAT locus. This defect was probably caused by transposition of a cassette originating from the hml alpha-2 allele by the process of the homothallic mating type switch. That the MAT locus of the nonmater cells is occupied by a DNA fragment indistinguishable from the Y alpha sequence in electrophoretic mobility was demonstrated by Southern hybridization of the EcoRI-HindIII fragment encoding the MAT locus with a cloned HML alpha gene as the probe. The hml alpha-2 mutation was revealed to be a one-base-pair deletion at the ninth base pair in the X region from the X and Y boundary of the HML locus. This mutation gave rise to a shift in the open reading frame of the alpha 2 cistron. A molecular mechanism for the mating type switch associated with the occurrence of sporogenous diploid cells in the mutant culture is discussed.

scholarly journals MALE-SPECIFIC LETHAL MUTATIONS OF DROSOPHILA MELANOGASTER. II. PARAMETERS OF GENE ACTION DURING MALE DEVELOPMENT

Genetics ◽  
1983 ◽  
Vol 105 (4) ◽  
pp. 881-896
Author(s):  
John M Belote
Keyword(s):  
Growth Patterns ◽  
Gene Products ◽  
Wild Type ◽  
Linked Gene ◽  
Lethal Mutations ◽  
Male Specific Lethal ◽  
A Cell ◽  
Type Gene ◽  
Diploid Cells ◽  
Male Specific

ABSTRACT The male-specific lethal mutations (msl's) identify loci whose wild-type gene products are essential for male, but not female, viability. Earlier studies in which X-linked gene activities were monitored in msl/msl male larvae demonstrated that these genes are responsible for setting and/or maintaining the level of X chromosome transcription in males (i.e., they are necessary for proper dosage compensation). The present study examines several important questions concerning their mode of action during development—The results of an examination of the effects of an msl-1 deficiency on male-lethal phase and female viability suggest that this mutation is an amorph, or a severe hypomorph. The effects of rendering a fly mutant for more than one male-lethal mutation were also examined. Multiply mutant flies were no more severely affected than singly mutant ones. A gynandromorph analysis revealed that the male-limited lethality associated with msl-2 has no single lethal focus. Somatic clones of homozygous msl-2 cells were initiated at various times during development by X-ray-induced mitotic recombination. An examination of the viability, growth patterns and morphology of marked clones demonstrated that: (1) msl-2  + acts in a cell autonomous manner, (2) msl-2  + function is required not only in larval (polytene) cells as was shown in previous work but is also needed in the diploid cells that give rise to adult structures, (3) the msl-2  + gene is needed fairly late in development and perhaps continuously, (4) the msl-2 mutation does not affect sexual differentiation.

Regulation of STA1 gene expression by MAT during the life cycle of Saccharomyces cerevisiae

1989 ◽  
Vol 9 (9) ◽  
pp. 3992-3998
Author(s):  
A M Dranginis
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mating Type ◽  
Yeast Cells ◽  
Specific Gene ◽  
Activity Levels ◽  
Sporulation Medium ◽  
Mrna Levels ◽  
Yeast Saccharomyces Cerevisiae ◽  
Type Locus ◽  
Diploid Cells

STA1 encodes a secreted glucoamylase of the yeast Saccharomyces cerevisiae var. diastaticus. Glucoamylase secretion is controlled by the mating type locus MAT; a and alpha haploid yeast cells secrete high levels of the enzyme, but a/alpha diploid cells produce undetectable amounts. It has been suggested that STA1 is regulated by MATa2 (I. Yamashita, Y. Takano, and S. Fukui, J. Bacteriol. 164:769-773, 1985), which is a MAT transcript of previously unknown function. In contrast, this work shows that deletion of the entire MATa2 gene had no effect on STA1 regulation but that deletion of MATa1 sequences completely abolished mating-type control. In all cases, glucoamylase activity levels reflected STA1 mRNA levels. It appears that STA1 is a haploid-specific gene that is regulated by MATa1 and a product of the MAT alpha locus and that this regulation occurs at the level of RNA accumulation. STA1 expression was also shown to be glucose repressible. STA1 mRNA was induced in diploids during sporulation along with SGA, a closely linked gene that encodes an intracellular sporulation-specific glucoamylase of S. cerevisiae. A diploid strain with a MATa1 deletion showed normal induction of STA1 in sporulation medium, but SGA expression was abolished. Therefore, these two homologous and closely linked glucoamylase genes are induced by different mechanisms during sporulation. STA1 induction may be a response to the starvation conditions necessary for sporulation, while SGA induction is governed by the pathway by which MAT regulates sporulation. The strain containing a complete deletion of MATa2 grew, mated, and sporulated normally.

scholarly journals Potential Repurposing of Known Drugs as Potent Bacterial β-Glucuronidase Inhibitors

2012 ◽  
Vol 17 (7) ◽  
pp. 957-965 ◽  
Author(s):  
Syed Ahmad ◽  
Mark A. Hughes ◽  
Li-An Yeh ◽  
John E. Scott
Keyword(s):  
Escherichia Coli ◽  
Gastrointestinal Tract ◽  
Active Metabolite ◽  
Enzyme Assay ◽  
Intestinal Bacteria ◽  
Tricyclic Antidepressant ◽  
Gus Activity ◽  
Significant Activity ◽  
A Cell ◽  
Approved Drugs

The active metabolite of the chemotherapeutic irinotecan, SN-38, is detoxified through glucuronidation and then excreted into the gastrointestinal tract. Intestinal bacteria convert the glucuronidated metabolite back to the toxic SN-38 using β-glucuronidase (GUS), resulting in debilitating diarrhea. Inhibiting GUS activity may relieve this side effect of irinotecan. In this study, we sought to determine whether any known drugs have GUS inhibitory activity. We screened a library of Food and Drug Administration–approved drugs with a cell-free biochemical enzyme assay using purified bacterial GUS. After triage, five drugs were confirmed to inhibit purified bacterial GUS. Three of these were the monoamine oxidase inhibitors nialamide, isocarboxazid, and phenelzine with average IC50 values for inhibiting GUS of 71, 128, and 2300 nM, respectively. The tricyclic antidepressant amoxapine (IC50 = 388 nM) and the antimalarial mefloquine (IC50 = 1.2 µM) also had activity. Nialamide, isocarboxazid, and amoxapine had no significant activity against purified mammalian GUS but showed potent activity for inhibiting endogenous GUS activity in a cell-based assay using living intact Escherichia coli with average IC50 values of 17, 336, and 119 nM, respectively. Thus, nialamide, isocarboxazid, and amoxapine have potential to be repurposed as therapeutics to reduce diarrhea associated with irinotecan chemotherapy and warrant further investigation for this use.

Biodiversity as a source of new pharmacophores: A new theory of memory III

2005 ◽  
Vol 77 (1) ◽  
pp. 75-81 ◽  
Author(s):  
◽  
M. Iqbal Choudhary
Keyword(s):  
Natural Products ◽  
Anticancer Agents ◽  
Neurological Disorders ◽  
In Vitro Bioassay ◽  
Natural Sources ◽  
Chemical Basis ◽  
Antioxidant Agents ◽  
A Cell ◽  
The Brain

Several classes of natural products with significant inhibitory activity against target enzymes involved in several diseases have been identified. Spectrophotometer and high-throughput assays were used to assess the inhibition of prolyl endopeptidase (PEP), which led us to some novel inhibitors having potential as anticancer agents. Inhibition of cholinesterase enzymes has led to the discovery of new inhibitors with potential for use in Alzheimer’s disease and other neurological disorders. We have also discovered several potent antioxidant agents from natural sources by using a battery of antioxidant assays. Anti-inflammatory activity of a number of natural products was assayed through a cell-based in vitro bioassay. This article also contains a section on a slightly different topic of chemical basis of memory as presented during the lecture. The theory of the chemical basis of memory based on hydrogen bonding in the brain is further elaborated.

scholarly journals “Modulation of apoptosis controls inhibitory interneuron number in the cortex”

2017 ◽  
Author(s):  
Myrto Denaxa ◽  
Guilherme Neves ◽  
Adam Rabinowitz ◽  
Sarah Kemlo ◽  
Petros Liodis ◽  
...  
Keyword(s):  
Physiological State ◽  
Inhibitory Interneuron ◽  
Projection Neurons ◽  
Activity Levels ◽  
Cortical Interneurons ◽  
A Cell ◽  
Cell Type Specific ◽  
Excitatory Projection ◽  
Cortical Excitation ◽  
The Right

AbstractCortical networks are composed of excitatory projection neurons and inhibitory interneurons. Finding the right balance between the two is important for controlling overall cortical excitation and network dynamics. However, it is unclear how the correct number of cortical interneurons (CIs) is established in the mammalian forebrain. CIs are generated in excess from basal forebrain progenitors and their final numbers are adjusted via an intrinsically determined program of apoptosis that takes place during an early postnatal window. Here, we provide evidence that the extent of CI apoptosis during this critical period is plastic, cell type specific and can be reduced in a cell autonomous manner by acute increases in neuronal activity. We propose that the physiological state of the emerging neural network controls the activity levels of local CIs to modulate their numbers in a homeostatic manner.

459 Validation of a cell panel for preclinical evaluation of antitumor efficacy and toxicity of anticancer agents

2010 ◽  
Vol 8 (7) ◽  
pp. 145
Author(s):  
C. Haglund ◽  
A. Åleskog ◽  
P. Nygren ◽  
J. Gullbo ◽  
M. Höglund ◽  
...  
Keyword(s):  
Anticancer Agents ◽  
Antitumor Efficacy ◽  
Preclinical Evaluation ◽  
A Cell

scholarly journals Targeting the Mitotic Catastrophe Signaling Pathway in Cancer

2015 ◽  
Vol 2015 ◽  
pp. 1-13 ◽  
Author(s):  
Margaret M. Mc Gee
Keyword(s):  
Molecular Mechanism ◽  
Cell Fate ◽  
Anticancer Agents ◽  
Therapeutic Benefit ◽  
Mitotic Catastrophe ◽  
Patient Response ◽  
Therapeutic Advantage ◽  
Prevention And Treatment ◽  
A Cell ◽  
Bona Fide

Mitotic catastrophe, as defined in 2012 by the International Nomenclature Committee on Cell Death, is abona fideintrinsic oncosuppressive mechanism that senses mitotic failure and responds by driving a cell to an irreversible antiproliferative fate of death or senescence. Thus, failed mitotic catastrophe can promote the unrestrained growth of defective cells, thereby representing a major gateway to tumour development. Furthermore, the activation of mitotic catastrophe offers significant therapeutic advantage which has been exploited in the action of conventional and targeted anticancer agents. Yet, despite its importance in tumour prevention and treatment, the molecular mechanism of mitotic catastrophe is not well understood. A better understanding of the signals that determine cell fate following failed or defective mitosis will reveal new opportunities to selectively target and enhance the programme for therapeutic benefit and reveal biomarkers to predict patient response. This review is focused on the molecular mechanism of mitotic catastrophe induction and signalling and highlights current strategies to exploit the process in cancer therapy.

scholarly journals Centrosome Clustering in the Development of Bovine Binucleate Trophoblast Giant Cells

2016 ◽  
Vol 203 (5) ◽  
pp. 287-294 ◽  
Author(s):  
Karl Klisch ◽  
Elisabeth M. Schraner ◽  
Alois Boos
Keyword(s):  
Cell Cycle ◽  
Tumor Cells ◽  
Molecular Mechanisms ◽  
Giant Cells ◽  
Cell Nuclei ◽  
Pericentriolar Material ◽  
A Cell ◽  
Pass Through ◽  
Diploid Cells ◽  
Trophoblast Giant Cells

Binucleate trophoblast giant cells (BNC) are the characteristic feature of the ruminant placenta. During their development, BNC pass through 2 acytokinetic mitoses and become binucleate with 2 tetraploid nuclei. In this study, we investigate the number and location of centrosomes in bovine BNC. Centrosomes typically consist of 2 centrioles surrounded by electron-dense pericentriolar material. Duplication of centrosomes is tightly linked to the cell cycle, which ensures that the number of centrosomes remains constant in proliferating diploid cells. Alterations of the cell cycle, which affect the number of chromosome sets, also affect the number of centrosomes. In this study, we use placentomal tissue from pregnant cows (gestational days 80-230) for immunohistochemical staining of γ-tubulin (n = 3) and transmission electron microscopy (n = 3). We show that mature BNC have 4 centrosomes with 8 centrioles, clustered in the angle between the 2 cell nuclei. During the second acytokinetic mitosis, the centrosomes must be clustered to form the poles of a bipolar spindle. In rare cases, centrosome clustering fails and tripolar mitosis leads to the formation of trinucleate “BNC”. Generally, centrosome clustering occurs in polyploid tumor cells, which have an increased number of centrioles, but it is absent in proliferating diploid cells. Thus, inhibition of centrosome clustering in tumor cells is a novel promising strategy for cancer treatment. BNC are a cell population in which centrosome clustering occurs as part of the normal life history. Thus, they might be a good model for the study of the molecular mechanisms of centrosome clustering.

scholarly journals Oncology Drug Discovery Applications Using the FMAT™ 8100 HTS System

2003 ◽  
Vol 8 (1) ◽  
pp. 81-88 ◽  
Author(s):  
Jennifer Y. Lee ◽  
Sheri Miraglia ◽  
Xiongwei Yan ◽  
Elana Swartzman ◽  
Susan Cornell-Kennon ◽  
...  
Keyword(s):  
Anticancer Agents ◽  
High Throughput Screening ◽  
Enzyme Assay ◽  
Annexin V ◽  
Enzyme Assays ◽  
Binding Assays ◽  
Src Tyrosine Kinase ◽  
Apoptosis Assay ◽  
Peptide Interaction ◽  
Homogeneous Assays

High-throughput screening (HTS) for potential anticancer agents requires a broad portfolio of assay platforms that may include kinase enzyme assays, protein-protein binding assays, and functional cell-based apoptosis assays. The authors have explored the use of fluorometric microvolume assay technology (the FMAT™ 8100 HTS System) in three distinct homogeneous HTS assays: (1) a Src tyrosine kinase enzyme assay, (2) a Grb2-SH2 protein-peptide interaction assay, and (3) an annexin V binding apoptosis assay. Data obtained from all three assays suggest that the FMAT system should facilitate the implementation of homogeneous assays for a wide variety of molecular targeted and cell-based screens. ( Journal of Biomolecular Screening 2003:81-88)

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