DNA ligation in relation to DNA strand breaks in phytohemagglutinin- stimulated blast cells from acute lymphoblastic and nonlymphoblastic leukemia

DNA ligase activity was determined in the WBCs from 306 cases of acute lymphoblastic leukemia (ALL) and acute nonlymphocytic leukemia (ANLL). In T-ALL cells this activity was either low or absent. DNA analysis by nucleoid, alkaline elution, and alkaline sucrose centrifugation after cells were embedded in agarose inserts has shown more DNA breaks in T- ALL than in ANLL blasts. Phytohemagglutinin stimulation of T-ALL blasts resulted in the apparent joining of the DNA breaks. Apparent identical results can be obtained by the incubation of DNA with exogenous DNA ligase. The authors suggest that this enzyme is a crucially regulated step of replication and subsequent proliferation in this type of leukemia.

Download Full-text

DNA ligation in relation to DNA strand breaks in phytohemagglutinin- stimulated blast cells from acute lymphoblastic and nonlymphoblastic leukemia

Blood ◽

10.1182/blood.v72.2.648.648 ◽

1988 ◽

Vol 72 (2) ◽

pp. 648-654

Author(s):

SA Feon ◽

RM Valerius ◽

NM Genetet ◽

I Bernard-Griffiths ◽

PY Le Prise ◽

...

Keyword(s):

Dna Analysis ◽

Lymphoblastic Leukemia ◽

Dna Ligase ◽

Dna Breaks ◽

Alkaline Elution ◽

Exogenous Dna ◽

Strand Breaks ◽

Dna Ligation ◽

Blast Cells ◽

Dna Strand

Abstract DNA ligase activity was determined in the WBCs from 306 cases of acute lymphoblastic leukemia (ALL) and acute nonlymphocytic leukemia (ANLL). In T-ALL cells this activity was either low or absent. DNA analysis by nucleoid, alkaline elution, and alkaline sucrose centrifugation after cells were embedded in agarose inserts has shown more DNA breaks in T- ALL than in ANLL blasts. Phytohemagglutinin stimulation of T-ALL blasts resulted in the apparent joining of the DNA breaks. Apparent identical results can be obtained by the incubation of DNA with exogenous DNA ligase. The authors suggest that this enzyme is a crucially regulated step of replication and subsequent proliferation in this type of leukemia.

Download Full-text

The effect of anoxia on anthracycline-induced DNA damage in the RPMI-6410 human lymphoblastoid cell line

Canadian Journal of Biochemistry ◽

10.1139/o82-111 ◽

1982 ◽

Vol 60 (9) ◽

pp. 873-876 ◽

Cited By ~ 1

Author(s):

L. Brox ◽

B. Gowans ◽

R. To ◽

A. Belch

Keyword(s):

Cell Line ◽

Lymphoblastoid Cell Line ◽

Dna Strand Breaks ◽

Primary Role ◽

Alkaline Elution ◽

Strand Breaks ◽

Human Lymphoblastoid Cell ◽

Strand Breakage ◽

Human Lymphoblastoid Cell Line ◽

Dna Strand

The alkaline elution procedure developed by Kohn and co-workers was used with the RPMI-6410 cultured human lymphoblastoid cell line to examine the hypothesis that anthracycline-induced DNA strand scission is mediated by oxygen- or superoxide-derived free radicals. Hypoxia was induced by gassing with nitrogen containing 5% carbon dioxide and less than 4 ppm oxygen. Alkaline elution studies showed hypoxia was induced, as the oxygen enhancement ratios for DNA strand breaks was 2.4 and 2.6 for the 250 R ± oxygen and the 500 R ± oxygen (1 R = 2.58 × 10−4 C/kg) experiments, respectively. The pattern of adriamycin-induced DNA strand breaks and cross-linking was not affected by hypoxia with 1-h adriamycin exposures between 0.05 and 1.0 μg/mL. Similarly, 1-h exposures of N-trifluoroacetyladriamycin-14-valerate at 3 or 10 μg/mL gave essentially identical alkaline elution profiles in the presence or absence of oxygen. These results do not support the hypothesis that oxygen-derived radicals play a primary role in anthracycline-induced DNA strand breakage.

Download Full-text

Clinical response to deoxycoformycin in chronic lymphoid neoplasms and biochemical changes in circulating malignant cells in vivo

Blood ◽

10.1182/blood.v72.6.1884.1884 ◽

1988 ◽

Vol 72 (6) ◽

pp. 1884-1890 ◽

Cited By ~ 13

Author(s):

AD Ho ◽

K Ganeshaguru ◽

WU Knauf ◽

G Dietz ◽

I Trede ◽

...

Keyword(s):

Clinical Response ◽

Dna Strand Breaks ◽

Biochemical Changes ◽

Leukemic Cells ◽

Malignant Cells ◽

Dna Breaks ◽

Strand Breaks ◽

Lymphoid Neoplasms ◽

Dna Strand

Abstract Deoxycoformycin (DCF), an adenosine deaminase (ADA) inhibitor, has been shown to be active in lymphoid neoplasms. The mechanism of cytotoxicity might involve accumulation of deoxyadenosine triphosphate (dATP), depletion of the nicotinamide adenine dinucleotide (NAD) and ATP pool, induction of double-stranded DNA strand breaks, or inhibition of S- adenosyl homocysteine hydrolase (SAH-hydrolase). We have investigated the biochemical changes in the circulating malignant cells of patients with chronic leukemia/lymphoma who were treated with DCF (4 mg/m2 weekly). Blood samples were taken from 17 patients with 60% or more circulating leukemic cells before, 4, 24, and 48 hours and five days after the first administration of DCF. Leukemic cells were separated and studied for changes in ADA, dATP, ATP, NAD, and SAH-hydrolase levels and DNA strand breaks and the data analyzed according to clinical response. Inhibition of ADA activity was found in all except one patient at 4 to 24 hours after the first administration of DCF. dATP started to accumulate at four hours, reached a maximum level between 24 and 48 hours, and returned to base values on the fifth day. Intracellular ATP and NAD levels were transiently reduced in some of the patients. However, no correlation between these changes and a clinical response could be found. DNA strand breaks could be studied in 13 patients. A significant increase in DNA breaks at 24 to 48 hours was found in six of the seven responders but only in one of the six nonresponders. At 24 hours, SAH-hydrolase levels were reduced in all seven responders studied, but only in two of the seven nonresponders. The difference in inhibition of SAH-hydrolase was statistically significant (P = .0023). These results suggest that DNA strand breaks and inhibition of SAH-hydrolase correlate with clinical response.

Download Full-text

Elevated Levels of DNA Strand Breaks Induced by a Base Analog in the Human Cell Line with the P32T ITPA Variant

Journal of Nucleic Acids ◽

10.4061/2010/872180 ◽

2010 ◽

Vol 2010 ◽

pp. 1-11 ◽

Cited By ~ 11

Author(s):

Irina S.-R. Waisertreiger ◽

Miriam R. Menezes ◽

James Randazzo ◽

Youri I. Pavlov

Keyword(s):

Cell Line ◽

Human Cell Line ◽

Dna Breaks ◽

Nucleotide Biosynthesis ◽

Purine Base ◽

Strand Breaks ◽

Inosine Triphosphate Pyrophosphatase ◽

Base Analog ◽

Deoxynucleoside Triphosphate ◽

Dna Strand

Base analogs are powerful antimetabolites and dangerous mutagens generated endogenously by oxidative stress, inflammation, and aberrant nucleotide biosynthesis. Human inosine triphosphate pyrophosphatase (ITPA) hydrolyzes triphosphates of noncanonical purine bases (i.e., ITP, dITP, XTP, dXTP, or their mimic: 6-hydroxyaminopurine (HAP) deoxynucleoside triphosphate) and thus regulates nucleotide pools and protects cells from DNA damage. We demonstrate that the model purine base analog HAP induces DNA breaks in human cells and leads to elevation of levels of ITPA. A human polymorphic allele of theITPA, 94C->A encodes for the enzyme with a P32T amino-acid change and leads to accumulation of nonhydrolyzed ITP. The polymorphism has been associated with adverse reaction to purine base-analog drugs. The level of both spontaneous and HAP-induced DNA breaks is elevated in the cell line with the ITPA P32T variant. The results suggested that human ITPA plays a pivotal role in the protection of DNA from noncanonical purine base analogs.

Download Full-text

The role of Sp1 in the detection and elimination of cells with persistent DNA strand breaks

NAR Cancer ◽

10.1093/narcan/zcaa004 ◽

2020 ◽

Vol 2 (2) ◽

Author(s):

Polina S Loshchenova ◽

Svetlana V Sergeeva ◽

Sally C Fletcher ◽

Grigory L Dianov

Keyword(s):

Dna Damage ◽

Cancer Therapy ◽

Genome Stability ◽

Dna Strand Breaks ◽

Dna Lesions ◽

Exogenous Dna ◽

Strand Breaks ◽

Specificity Factor ◽

Dna Strand

Abstract Maintenance of genome stability suppresses cancer and other human diseases and is critical for organism survival. Inevitably, during a life span, multiple DNA lesions can arise due to the inherent instability of DNA molecules or due to endogenous or exogenous DNA damaging factors. To avoid malignant transformation of cells with damaged DNA, multiple mechanisms have evolved to repair DNA or to detect and eradicate cells accumulating unrepaired DNA damage. In this review, we discuss recent findings on the role of Sp1 (specificity factor 1) in the detection and elimination of cells accumulating persistent DNA strand breaks. We also discuss how this mechanism may contribute to the maintenance of physiological populations of healthy cells in an organism, thus preventing cancer formation, and the possible application of these findings in cancer therapy.

Download Full-text

Mechanistic insight into the role of Poly(ADP-ribosyl)ation in DNA topology modulation and response to DNA damage

Mutagenesis ◽

10.1093/mutage/gez045 ◽

2019 ◽

Vol 35 (1) ◽

pp. 107-118

Author(s):

Bakhyt T Matkarimov ◽

Dmitry O Zharkov ◽

Murat K Saparbaev

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Genotoxic Stress ◽

Strand Break ◽

Dna Supercoiling ◽

Dna Strand Breaks ◽

Chromosomal Dna ◽

Dna Breaks ◽

Strand Breaks ◽

Dna Strand

Abstract Genotoxic stress generates single- and double-strand DNA breaks either through direct damage by reactive oxygen species or as intermediates of DNA repair. Failure to detect and repair DNA strand breaks leads to deleterious consequences such as chromosomal aberrations, genomic instability and cell death. DNA strand breaks disrupt the superhelical state of cellular DNA, which further disturbs the chromatin architecture and gene activity regulation. Proteins from the poly(ADP-ribose) polymerase (PARP) family, such as PARP1 and PARP2, use NAD+ as a substrate to catalyse the synthesis of polymeric chains consisting of ADP-ribose units covalently attached to an acceptor molecule. PARP1 and PARP2 are regarded as DNA damage sensors that, upon activation by strand breaks, poly(ADP-ribosyl)ate themselves and nuclear acceptor proteins. Noteworthy, the regularly branched structure of poly(ADP-ribose) polymer suggests that the mechanism of its synthesis may involve circular movement of PARP1 around the DNA helix, with a branching point in PAR corresponding to one complete 360° turn. We propose that PARP1 stays bound to a DNA strand break end, but rotates around the helix displaced by the growing poly(ADP-ribose) chain, and that this rotation could introduce positive supercoils into damaged chromosomal DNA. This topology modulation would enable nucleosome displacement and chromatin decondensation around the lesion site, facilitating the access of DNA repair proteins or transcription factors. PARP1-mediated DNA supercoiling can be transmitted over long distances, resulting in changes in the high-order chromatin structures. The available structures of PARP1 are consistent with the strand break-induced PAR synthesis as a driving force for PARP1 rotation around the DNA axis.

Download Full-text

Mouse Strains with an Active H2-Ea Meiotic Recombination Hot Spot Exhibit Increased Levels of H2-Ea-Specific DNA Breaks in Testicular Germ Cells

Molecular and Cellular Biology ◽

10.1128/mcb.24.4.1655-1666.2004 ◽

2004 ◽

Vol 24 (4) ◽

pp. 1655-1666 ◽

Cited By ~ 28

Author(s):

Jian Qin ◽

Laura L. Richardson ◽

Maria Jasin ◽

Mary Ann Handel ◽

Norman Arnheim

Keyword(s):

Germ Cells ◽

Hot Spot ◽

Cell Types ◽

Mouse Strains ◽

Hydroxyl Groups ◽

Dna Breaks ◽

Strand Breaks ◽

Dna Strand ◽

Pachytene Spermatocytes ◽

Old Mice

ABSTRACT We devised a sensitive method for the site-specific detection of rare meiotic DNA strand breaks in germ cell-enriched testicular cell populations from mice that possess or lack an active recombination hot spot at the H2-Ea gene. Using germ cells from adult animals, we found an excellent correlation between the frequency of DNA breaks in the 418-bp H2-Ea hot spot and crossover activity. The temporal appearance of DNA breaks was also studied in 7- to 18-day-old mice with an active hot spot during the first waves of spermatogenesis. The number of DNA breaks detected rose as leptotene and zygotene spermatocytes populate the testis with a peak at day 14 postpartum, when leptotene, zygotene, and early pachytene spermatocytes are the most common meiotic prophase I cell types. The number of DNA breaks drops precipitously 1 day later, when middle to late pachytene spermatocytes become the dominant subtype. The recombination-related breaks in the hot spot likely reflect SPO11-induced double-strand breaks and/or recombination intermediates containing free 3′ hydroxyl groups.

Download Full-text

Contrasting patterns of DNA strand breakage and ADP-ribosylation- dependent DNA ligation during granulocyte and monocyte differentiation

Blood ◽

10.1182/blood.v69.4.1114.bloodjournal6941114 ◽

1987 ◽

Vol 69 (4) ◽

pp. 1114-1119 ◽

Cited By ~ 1

Author(s):

Z Khan ◽

GE Francis

Keyword(s):

Structural Changes ◽

Dna Supercoiling ◽

Monocytic Differentiation ◽

Dna Breaks ◽

Strand Breaks ◽

Colony Stimulating Activity ◽

Strand Breakage ◽

Macrophage Colony ◽

Dna Strand ◽

Dna Strand Breakage

Previous studies have shown that structural changes in DNA, including the ligation of pre-existing DNA breaks and the opening and closure of new breaks, occur shortly after exposure of granulomonocytic precursors (CFU-GM) to granulocyte-macrophage colony stimulating activity (GM- CSA). Monocytic differentiation of CFU-GM is selectively inhibited by compounds known to inhibit the nuclear enzyme ADP-ribosyl transferase (ADPRT). Since this enzyme, which transfers ADP-ribose units to chromatin proteins, is known to activate DNA ligase, we attempted to determine whether ligation of one or both types of DNA break is required for monocytic differentiation. Breaks in DNA were examined using the nucleoid sedimentation technique in which DNA breaks cause loss of DNA supercoiling in nucleoids and concomitant changes in their sedimentation through neutral sucrose gradients. We here report that two distinct patterns of DNA strand breakage and ligation are associated with differentiation to the granulocyte and monocyte lineages. Monocytic inducers (phorbolester and vitamin D3) predominantly produce closure of pre-existing strand breaks, whereas granulocytic inducers (granulocyte colony stimulating activity, G-CSA; retinoic acid) cause opening and closure of new breaks. Only ligation of the pre-existing breaks is highly sensitive to inhibition by 3- methoxybenzamide (a potent ADPRT inhibitor), and only monocytic differentiation is impaired by addition of this compound. These findings suggest that DNA structural changes may be directly involved in granulocyte-macrophage switching.

Download Full-text

Induction of CAF-1 Expression in Response to DNA Strand Breaks in Quiescent Human Cells

Molecular and Cellular Biology ◽

10.1128/mcb.26.5.1839-1849.2006 ◽

2006 ◽

Vol 26 (5) ◽

pp. 1839-1849 ◽

Cited By ~ 30

Author(s):

Arman Nabatiyan ◽

Dávid Szüts ◽

Torsten Krude

Keyword(s):

Excision Repair ◽

Dna Strand Breaks ◽

Significant Loss ◽

Human Cells ◽

Dna Breaks ◽

Strand Breaks ◽

Quiescent Cells ◽

Dna Strand

ABSTRACT Genome stability in eukaryotic cells is maintained through efficient DNA damage repair pathways, which have to access and utilize chromatin as their natural template. Here we investigate the role of chromatin assembly factor 1 (CAF-1) and its interacting protein, PCNA, in the response of quiescent human cells to DNA double-strand breaks (DSBs). The expression of CAF-1 and PCNA is dramatically induced in quiescent cells upon the generation of DSBs by the radiomimetic drug bleocin (a bleomycin compound) or by ionizing radiation. This induction depends on DNA-PK. CAF-1 and PCNA are recruited to damaged chromatin undergoing DNA repair of single- and double-strand DNA breaks by the base excision repair and nonhomologous end-joining pathways, respectively, in the absence of extensive DNA synthesis. CAF-1 prepared from repair-proficient quiescent cells after induction by bleocin mediates nucleosome assembly in vitro. Depletion of CAF-1 by RNA interference in bleocin-treated quiescent cells in vivo results in a significant loss of cell viability and an accumulation of DSBs. These results support a novel and essential role for CAF-1 in the response of quiescent human cells to DSBs, possibly by reassembling chromatin following repair of DNA strand breaks.

Download Full-text