Variations in some molecular events during the early phases of the Reuber H35 hepatoma cell cycle: III. Role of protein synthesis in the initiation of DNA synthesis and the mechanism of stimulation of protein synthesis by serum

Abstract

Induction of DNA synthesis by serum and amino acids has been investigated in cultured Reuber H35 hepatoma cells. Commitment of DNA synthesis was found to occur 6-8 hours before the actual start of this synthesis. The rate of initiation of DNA synthesis is proportional to the stimulation of protein synthesis by serum and/or amino acids. The increased protein synthesis is important for the proliferation only during the early period after serum addition. The withdrawal of serum and the inhibition by cycloheximide confirm this finding. Actinomycin D hardly influenced the early effect of serum on protein synthesis and it is concluded that the serum-stimulated protein synthesis is carried out on pre-existing mRNA's. The mechanism of stimulation of protein synthesis by serum has been studied by determination of the polyribosome size, the number of growing polypeptide chains, and the ribosomal transit time. The rate of the initiation of translation has been found to be specifically enhanced while the rate of elongation remained unchanged. Two-dimensional gel electrophoresis showed that the early stimulation of protein synthesis by serum involves all types of major cellular proteins, and no new proteins could be detected.