Free Ribosomes in Physiologically Nondividing Cells

Ribosomes of physiologically nondividing human peripheral blood lymphocytes were studied by ultraviolet absorbance measurements of cytoplasmic extracts subjected to ultracentrifugation in sucrose gradients at high and low ionic strength. At least 70% of the total cytoplasmic ribosomes were free ribosomes which sedimented at 80 S in low salt and dissociated to 40 S and 60 S subunits in high salt. These particles were not involved in protein synthesis. The remaining ribosomes were equally divided among native subunits, active monosomes, and larger polysomes. Free ribosomes were shown to exist as 80 S particles in the intact cell, and labeling studies indicated that they did not freely return to the pools of protein-synthesizing components. New ribosomes appeared first as native subunits and in polysomes. After a lag of 15 to 20 min, the particles began to enter the free ribosome pool. Thus, free ribosomes arise in the resting cell by a unidirectional flow which continuously removes particles from the protein-synthesizing pool and sequesters them as an accumulation of inactive 80 S particles. The transition from native subunits to free ribosomes is accompanied by functional changes in association behavior of subunits and by alteration of sedimentation behavior of the subunits. These changes may be due to absence of a protein or proteins from the free ribosomes which is required to permit effective dissociation of subunits prior to initiation of translation. Deficiency of this dissociation factor may be responsible for the continuous formation of free ribosomes in resting cells. Our data also suggest a limitation of the rate of initiation of protein synthesis which may result from deficiency of an initiation factor.