The Control of Ribosomal RNA Transcription in Lymphocytes. Evidence that the Rate of Chain Elongation Is the Limiting Factor

Abstract

During activation of lymphocytes by phytohaemagglutinin, the nuclear activity of RNA polymerase I increases with no proportional increase in the amount or catalytic efficiency of the enzyme in the cell. The mechanism by which rRNA transcription in lymphocytes is modified by phytohaemagglutinin stimulation was investigated. The following results were obtained. (a) In resting lymphocytes all RNA polymerase I1 molecules are bound to the template while a pool of excess free RNA polymerase I, not engaged in transcription, can be detected by its ability to transcribe added poly[d(A-T)]. (b) Although the free RNA polymerase I activity increases twofold to threefold during phytohaemagglutinin stimulation, there is no evidence that the free enzymes ever become engaged in transcription. (c) Most of the RNA chains in elongation in nuclei from resting lymphocytes are being elongated by class 11 RNA polymerase and their rate of elongation is much higher than that of other RNA species. (d) The same number of pre-rRNA chains are in the process of being elongated in nuclei from resting and stimulated lymphocytes. However, the rate of elongation of pre-rRNA, which is slow relative to the average in resting lymphocytes, increases twofold to threefold within 6 h of phytohaemagglutinin stimulation and rises to sixfold by 19 h. These results suggest that the control of rRNA transcription in phytohaemagglutininstimulated lymphocytes lies in the elongation step of transcription rather than in initiation, and that little or no additional rRNA template is transcribed in phytohaemagglutinin-stimulated lymphocytes.Addition of phytohaemagglutinin to a culture of peripheral blood lymphocytes stimulates a transition from the resting to the growing state. Measurements of the rates of RNA synthesis in isolated nuclei show a doubling of the activity of RNA polymerase 1 (pre-rRNA synthesis) as early as 4 h after the addition of phytohaemagglutinin. This increase rises to fivefold or sixfold by 20 h. The activity of RNA polymerase I1 (pre-mRNA synthesis) does not change during the first 12 h following phytohaemagglutinin stimulation and shows a twofold increase by 20 h [l]. Extraction of the polymerases from the lymphocytes and assay of their activity by using exogenous templates show that the increase in the nuclear activity of RNA polymerase I1 coincides with a proportional change in the amount or catalytic efficiency of the enzyme in the cells [2]. The level of polymerase I activity which can be extracted from the lymphocytes also increases following phytohaemagglutinin stimulation but not until after 12 h [2,3], and the magnitude of this increase is insufficient to account for the large alteration in the nuclear activity of the enzyme.Alterations in nuclear RNA polymerase activities with no proportional changes in the amount or catalytic efficiency of the enzymes in the cell have been reported for several physiological transitions (for review, see [4]) and it is clear that factors other than enzyme levels must determine the nuclear activity o...

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