Synthesis of polyadenylated messenger RNA during the cell cycle of Saccharomyces cerevisiae

Abstract

The rates of ribosomal and polyadenylated messenger RNA synthesis were measured through the cell cycle of the budding yeast Saccharomyces cerevisiae. An exponentially growing culture was pulse-labelled with [2- 3H]adenine for 0.1 generation time, then fractionated according to cell volume by zonal centrifugation. A control experiment with a synchronously dividing culture showed that zonal fractionation corresponded to a fractionation by stage in the cell cycle. Nucleic acids were extracted from each fraction of the cell cycle from the zonal rotor, and DNA, ribosomal and polyadenylated messenger RNAs were isolated. DNA synthesis occurred between 0.4 and 0.8 of the cell cycle as fractionated on the zonal rotor. The rate of ribosomal RNA synthesis per cell was constant during G 1‡ doubled early in the period of DNA synthesis, then remained constant at the new level for the remainder of the cell cycle. The rate of processing of ribosomal RNA precursors to mature ribosomal RNAs declined at the time of the increase in rate of ribosomal RNA synthesis, but recovered later in the cycle. This could mean that the pool of ribosomal proteins is small, and that the rate of ribosomal protein synthesis increases later in the cell cycle than the rate of ribosomal RNA synthesis. The rate of synthesis of polyadenylated messenger RNA was constant during G 1 and the earliest part of the period of DNA synthesis, then doubled over the remainder of the period of DNA synthesis. After DNA synthesis, the rate of polyadenylated messenger RNA synthesis remained steady for the remainder of the cell cycle. It is suggested that gene number controls the rates of ribosomal and messenger RNA synthesis. When the number of genes doubles during the DNA synthesis period, the amount of transcription also quickly doubles. Repeat experiments suggested that the genes coding for ribosomal RNA are reproducibly replicated earlier, and over a shorter proportion of the DNA replication period than the genes coding for messenger RNAs. It is calculated that a minimum of 625 genes coding for messenger RNA per haploid genome, transcribed at the maximum rate throughout the cell cycle, would be required to give the observed cell-cycle pattern of messenger RNA synthesis. The total content of polyadenylated messenger RNA of cells at different stages of the cell cycle was measured by the formation of ribonuclease-resistant complexes between polyadenylic acid sequences in total UNA from non-radioactive yeast and [ 3H]polyuridylic acid. The amount of polyadenylated messenger RNA per cell rose after the doubling of the rate of polyadenylated messenger RNA synthesis, and by the end of the cell cycle was twice the level at the beginning of the cycle. The cell-cycle patterns of messenger and ribosomal RNA synthesis are discussed in relation to the control of balanced growth and doubling of all cellular components during the cell cycle, and in relation to known patterns of enzyme activity during the cell cycle of budding yeast.