Calculation of absolute rates of RNA synthesis, accumulation, and degradation in tobacco callus in vivo.

Abstract

Uptake and incorporation data were used to calculate an absolute rate of ribonucleic acid (RNA) synthesis by assuming that incorporation of radioactive nucleosides into RNA was a first-order function of the nucleoside triphosphate cellular pool. Homogenized tissue was separated into an RNA and cellular pool fraction. Methods were developed to directly measure the specific radioactivity of specific nucleosides in each fraction. Exogenous [3H]uridine was taken up by the cells, converted to I[3H]uridine triphosphate ([3H]UTP) and [3H]cytidine triphosphate ([3H]CTP), and incorporated into RNA. All of the radioactivity in RNA was contained in either uridine or cytidine. The average rate of RNA synthesis (20 mg of RNA (g of RNA)-1h-1) was calculated from changes in the specific radioactivity of uridine or cytidine in RNA and UTP or CTP in the pool. Since ribosomal RNA does not turn over in exponentially growing tissue, its rate of synthesis was measured (14 mg g-1h-1) and compared with the rate of accumulation of RNA (12 mg g-1h-1). The similarity between ribosomal RNA synthesis and total RNA accumulation confirmed the assumption that these methods measured an absolute rate of RNA synthesis.