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RNA Polymerase Isolated from Bovine Lymphosarcoma by Sequential Low- and High-Salt Extraction¹

Departments of Biochemistry [A. A. K.] and Pathology [G. E. A., S. M. M., J. J. F.], School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19174

Bovine lymphosarcoma tissue has been extracted with low- and high-salt buffers [0.05 M Tris-Cl ± 0.3 M (NH₄)₂SO₄]. Diethylaminoethyl-Sephadex chromatography of both the high-salt and low-salt extracts yields RNA polymerases I and II, although low-salt extraction releases only one-third as much activity. Extraction by high salt of the residue from the low-salt extract, followed by diethylaminoethyl-Sephadex chromatography, yields additional enzyme activity with properties of Form II. Purification of the low-salt extract by protamine precipitation, elution with sodium succinate, and phosphocellulose chromatography yields a preparation of RNA polymerase (RNAP) with hybrid properties, combining the salt optimum of Form I, diethylaminoethyl-Sephadex elution pattern of Form II, and -amanitin sensitivity of Form III. RNAP_L transcribes native DNA and chromatin efficiently. More RNAP_L is recovered from lymphosarcoma tissue than from calf thymus.

¹ This investigation was supported by Grant 2-R01-GM-10390 from the NIH, USPHS. Support for maintenance of the herd with a high incidence of lymphosarcoma came from USPHS Cancer Center Grant 1-P01-CA-14193-01, USPHS Contract PH-43-6S-1013 within the Virus Cancer Program of the National Cancer Institute, and USPHS Grant 5-PO-6-RR-00182.

² Supported by Medical Scientist Training Program NIH Training Grant 5T05-GM-02046.

³ Supported by a Pennsylvania Plan Fellowship. Present address: Department of Pathology, School of Medicine, U.C.L.A. Center for Health Sciences, Los Angeles, Calif. 90024.

Received 8/13/74. Accepted 10/14/74.