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Studies on the Incorporation of L-Ethionine-ethyl-1-¹⁴C into the Transfer RNA of Rat Liver¹

Biology Division, NCI-AEC Carcinogenesis Program, Oak Ridge National Laboratory, Oak Ridge, Tennessee 37830

When rats were injected with two 50-µc doses of L-ethionine-ethyl-1-¹⁴C 24 hours apart, the mixed transfer RNA (tRNA) fraction was labeled to the extent of 200 cpm/A₂₆₀, but the radioactivity remaining after treatment of the DNA to remove RNA and protein was not significant. Ribosomal RNA, isolated by methylated albumin-kieselguhr chromatography, showed only 5 cpm/A₂₆₀.

The labeling of tRNA increased linearly from 1 to 12 hours following a single injection of L-ethionine-ethyl-1-¹⁴C, reached a maximum at 24 hours, and decayed with an apparent half-life of 2.5 days. When the labeling of tRNA was determined for several rat tissues, the liver tRNA was highest in both specific and total radioactivity.

The ethylation of tRNA by L-ethionine-ethyl-1-¹⁴C does not appear to proceed in a manner similar to normal methylation because S-adenosylethionine does not seem to be the active intermediate. This is suggested by the following results:

(a) The S-adenosylethionine content of the liver is the same or greater at 24 hours, when no tRNA ethylation can be seen, than it is at 3 hours, when tRNA ethylation is proceeding at a maximum rate.

(b) An in vitro system which functions well for the methylation of tRNA with S-adenosylmethionine does not ethylate tRNA with S-adenosylethionine.

(c) The presence of a 10-fold excess of methionine-¹²C increases the labeling of tRNA by L-ethionine-ethyl-1-¹⁴C even though it has been reported to inhibit the formation of S-adenosylethionine.
Also, levels of methionine-¹²C which prevent the formation of S-adenosylethionine completely, do not completely inhibit the ethylation of tRNA.

(d) The distribution of radioactivity across an RPC 2 chromatogram of tRNA labeled with ethionine-ethyl-1-¹⁴C is markedly different from that labeled with methionine-methyl-¹⁴C.

¹ Research jointly sponsored by the National Cancer Institute and by the U. S. Atomic Energy Commission under contract with the Union Carbide Corporation.

Received 10/23/67. Accepted 9/13/68.