Transcription of DNAs of Known Sequence after Injection into the Eggs and Oocytes of Xenopus laevis

Abstract

When the synthetic polynucleotide, poly[d(A‐T) · d(A‐T)] is injected into the eggs and oocytes of Xenopus laevis, a stimulation of RNA synthesis results. Analysis of extracted RNA by high‐voltage electrophoresis, shows that this stimulation of RNA synthesis is due to the production of poly[r(A‐U)] transcripts. The rate of poly[r(A‐U)] synthesis is calculated to be at least ten‐fold greater in eggs than in oocytes. The amount of poly[r(A‐U)] detectable in injected eggs has reached a maximum by 1.5 h after injection; in oocytes, however, poly[r(A‐U)] continues to accumulate between the third and 16th hour after injection. The transcripts range in length from less than 80 nucleotides up to over 2000 nucleotides long. The co‐injection of poly[d(A‐T)] · d(A‐T)] and α‐amanitin into oocytes, has shown that the synthesis of poly[r(A‐U)] is approximately 90% inhibited at a concentration of α‐amanitin which has no effect on the capacity of the oocyte to synthesize ribosomal and 4‐S RNA; thus the nucleoplasmic RNA polymerases IIa and/or IIb, are implicated as playing a major role in poly[r(A‐U)] synthesis in oocytes. When poly(dG) · poly(dC), poly(dA), poly(dA) · poly(dT) and poly[d(I‐C) · d(I‐C)] are individually injected into eggs only poly[d(I‐C) · d(I‐C)] is transcribed as efficiently as poly[d(A‐T) · d(A‐T)]. When calf thymus native or denatured DNA, polyoma, T2, T4 and φX DNAs are individually injected into eggs only the injection of calf thymus native DNA causes a detectable stimulation of RNA synthesis. The activities of crude preparations of egg and oocyte RNA polymerases are tested in vitro with different DNA templates. In contrast to the situation in vivo, it is found that poly[d(A‐T) · d(A‐T)] is as efficiently transcribed in vitro by oocyte polymerase as by egg polymerase. Additionally poly[d(A‐T) · d(A‐T)] is transcribed ten‐fold more efficiently in vitro than calf thymus native DNA. When poly(dA) · poly(dT), poly(dA), φX, T2, and calf thymus denatured DNA are tested in vitro, only calf thymus denatured DNA is transcribed to a significant extent. The above results are discussed in relation to the known synthetic activities of Xenopus eggs and oocytes. Copyright © 1975, Wiley Blackwell. All rights reserved