Measurement of terminal deoxynucleotidyl transferase mrna in clinical samples: A new parameter in analysis of leukemia cells

Wolf, Susan C.; Steinherz, Peter G.; Landau, Nathaniel R.; Silverstone, Allen E.

doi:10.1002/ajh.2830250305

Cited by 5 publications

(2 citation statements)

References 36 publications

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“…Autoradiograms were quantified with the above-described Bio-Rad Video Densitometer and computer program. Values obtained from the oligo-dT hybridization were used to normalize the TdT readings as described (Wolf et al, 1987). Multiple control samples had a normalized TdT/dA ratio within 10%.…”

Section: Protein Immunoprecipitation and Sds-page Electrophoresismentioning

confidence: 99%

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Phorbol ester regulation of terminal deoxynucleotidyl transferase, proliferation, and TcR α in a pre‐T cell line

Béland

Yuille

Hugunin

et al. 1990

Journal Cellular Physiology

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Terminal deoxynucleotidyl transferase (TdT) is a template-independent DNA polymerase that is transiently expressed during the normal development of T and B lymphocytes. Phorbol 12-myristate 13-acetate (PMA) has been reported to induce maturation-like changes, including the loss of TdT, in many leukemic cell lines. We investigated the mechanism of TdT repression by PMA in an early thymocyte-like cell line, RPMI 8402. At a concentration of 8 nM, PMA caused both repression of TdT synthesis and arrest of proliferation. At greater concentrations of PMA, these same changes initially occurred, but then cell proliferation resumed, and TdT was reexpressed. At both 8 and 160 nM PMA, TdT biosynthesis and TdT mRNA became undetectable within 8 hours, while cell proliferation and DNA synthesis were not significantly reduced until 16 hours. Growth arrest induced by serum starvation did not result in a similar reduction of TdT RNA even after 48 hours. With 160 nM PMA, TdT mRNA could be detected again by 24 hours, and proliferation resumed. Transcription run-off assays indicated that TdT RNA synthesis ceased within 1 hour after exposure to both 8 and 160 nM PMA. T cell receptor alpha (TcR alpha) RNA was induced when TdT RNA was repressed. TcR beta RNA levels were unchanged, and TcR gamma RNA was up-regulated. TdT gene repression and modulation of cell proliferation as well as induction of TcR gene expression are normal events during intrathymic T cell maturation. This cell model provides a system for analyzing the molecular regulation of these significant developmental events.

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Section: Protein Immunoprecipitation and Sds-page Electrophoresismentioning

confidence: 99%

“…'TdT RNA was measured as described. Values Indicated are the ratios of the area of the slots hybridized with TdT probe 317-2 to the area detected after rehyhridizing with end-labeled poly-dT (as a measure of total poly-A RNA) (Wolf et al, 1987). The above values are derived from analysis of several dilutions on one membrane.…”

Section: Pma and Gene Expressionmentioning

confidence: 99%

Phorbol ester regulation of terminal deoxynucleotidyl transferase, proliferation, and TcR α in a pre‐T cell line

Béland

Yuille

Hugunin

et al. 1990

Journal Cellular Physiology

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The Use of Non‐natural Nucleotides to Probe Template‐Independent DNA Synthesis

Berdis

McCutcheon

2007

ChemBioChem

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The vast majority of DNA polymerases use the complementary templating strand of DNA to guide each nucleotide incorporation. There are instances, however, in which polymerases can efficiently incorporate nucleotides in the absence of templating information. This process, known as translesion DNA synthesis, can alter the proper genetic code of an organism. To further elucidate the mechanism of template-independent DNA synthesis, we monitored the incorporation of various nucleotides at the "blunt-end" of duplex DNA by the high-fidelity bacteriophage T4 DNA polymerase. Although natural nucleotides are not incorporated at the blunt-end, a limited subset of non-natural indolyl analogues containing extensive pi-electron surface areas are efficiently utilized by the T4 DNA polymerase. These analogues possess high binding affinities that are remarkably similar to those measured during incorporation opposite an abasic site. In contrast, the k(pol) values are significantly lower during blunt-end extension when compared to incorporation opposite an abasic site. These kinetic differences suggest that the single-stranded region of the DNA template plays an important role during polymerization through stacking interactions with downstream bases, interactions with key amino acid residues, or both. In addition, we demonstrate that terminal deoxynucleotide transferase, a template-independent enzyme, can efficiently incorporate many of these non-natural nucleotides. However, that this unique polymerase cannot extend large, bulky non-natural nucleotides suggests that elongation is limited by steric constraints imposed by structural features present within the polymerase. Regardless, the kinetic data obtained from using either DNA polymerase indicate that template-independent synthesis can occur without the contributions of hydrogen-bonding interactions and suggest that pi-electron interactions play an important role in polymerization efficiency when templating information is not present.

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Gene Expression of Terminal Deoxynucleotidyl Transferase in Neoplastic Cells of Leukemia and Lymphoma

Oiwa

Koiwai

Kaneda

1989

Japanese Journal of Cancer Research

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Expression levels of terminal deoxynucleotidyl transferase (TdT) mRNA in fresh leukemia and lymphoma cells were measured by northern blotting analysis. Bands of 2.1 kb mRNA were detected in all of eight cases of TdT activity‐positive leukemias: two cases of null‐cell acute lymphoblastic leukemia (null‐ALL), two of common ALL, one of pre‐B ALL, one of T‐ALL, and two of chronic myelogenous leukemia in blastic crisis. One of the null‐ALL and one of the common ALL cases also showed large TdT mRNA (3.3 kb). Since all TdT activity‐positive samples exhibited TdT mRNA, the TdT gene might be mainly regulated at the transcription level in leukemic cells. An elevated level of 2.1 kb TdT mRNA was also detected in one lymphoma case, where neither TdT activity nor immunoreactive TdT was detected. The extensive chromosomal abnormality demonstrated in this case might be associated with the translational anomaly of TdT.

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Measurement of terminal deoxynucleotidyl transferase mrna in clinical samples: A new parameter in analysis of leukemia cells

Cited by 5 publications

References 36 publications

Phorbol ester regulation of terminal deoxynucleotidyl transferase, proliferation, and TcR α in a pre‐T cell line

Phorbol ester regulation of terminal deoxynucleotidyl transferase, proliferation, and TcR α in a pre‐T cell line

The Use of Non‐natural Nucleotides to Probe Template‐Independent DNA Synthesis

Gene Expression of Terminal Deoxynucleotidyl Transferase in Neoplastic Cells of Leukemia and Lymphoma

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