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Poly(2'-O-methylcytidylate)-directed DNA Polymerase and Terminal Deoxynucleotidyltransferase Activities in Particulate Fractions Obtained from Human Spleens¹

Laboratory of Molecular Virology, Memorial Sloan-Kettering Cancer Center, New York, New York 10021

We have modified a procedure involving chromatography on polycytidylate-agarose and the template-primer complex poly(2'-O-methylcytidylate)·oligodeoxyguanylate which we have shown to be useful in the detection and partial purification of two forms of reverse transcriptase from extracts of retrovirus-infected cells for the detection of poly(2'-O-methylcytidylate)-directed DNA polymerase activity in particulate fractions obtained from the involved spleens of patients with leukemia, lymphoma, and other hematopoietic disorders. Isopyknically banded particulate fractions from human spleens could not be assayed directly, inasmuch as they were found to inhibit murine retroviral DNA polymerase activity. Assays for the detection of the primer-dependent, template-independent DNA polymerase, terminal deoxynucleotidyltransferase (TdT) were also performed. Poly(2'-O-methylcytidylate)-directed DNA polymerase activity was identified in 6 of 16 spleen samples from patients with leukemia and in 4 of 10 spleen samples from patients with lymphomas. A similar polymerase activity was identified in particulate fractions in 1 of 5 spleen samples from patients free of cancer. In one instance, activity was sufficient to show that oligodeoxythymidylate-primed globin messenger RNA could apparently direct DNA synthesis by polymerase-containing fractions. When present, poly(2'-O-methylcytidylate)-directed DNA polymerase activity was found in sucrose density gradient fractions similar in density to retroviruses or to retrovirus "core" particles.

TdT activity was found in particulate fractions in 3 of 16 spleen samples from patients with leukemia and in 3 of 10 spleen samples from patients with lymphoma. TdT was also found in 1 of 5 spleen samples from cancer-free patients. The presence of TdT activity in such particulate fractions suggests caution in the interpretation of DNA polymerase assay results using hematopoietic tissue unless sufficient controls for the detection of this enzyme are provided. The unexpected presence of TdT in spleen samples may relate to its preferential concentration by the methods used, a possible tendency for TdT to associate through hydrophobic interaction with microsomal material or the ecotaxopathy and sequestration of certain quantities of TdT-containing cells in spleen for as yet unknown reasons.

¹ This research was supported in part by USPHS Research Grants CA-08748, CA-18369, and CA-20194.

² To whom requests for reprints should be addressed.

Received 10/ 1/79. Accepted 12/21/79.