The fate of membrane-bound ribosomes following the termination of protein synthesis

Contemporary models for protein translocation in the mammalian endoplasmic reticulum (ER) identify the termination of protein synthesis as the signal for ribosome release from the ER membrane, We have utilized morphometric an d biochemical methods to assess directly the fate of membrane-bound ribosom es following the termination of protein synthesis. In these studies, tissue culture cells were treated with cycloheximide to inhibit elongation, with pactamycin to inhibit initiation, or with puromycin to induce premature cha in termination, and ribosome-membrane interactions were subsequently analyz ed. It was found that following the termination of protein synthesis, the m ajority of ribosomal particles remained membrane-associated. Analysis of th e subunit structure of the membrane-bound ribosomal particles remaining aft er termination was conducted by negative stain electron microscopy and sucr ose gradient sedimentation. By both methods of analysis, the termination of protein synthesis on membrane-bound ribosomes was accompanied by the relea se of small ribosomal subunits from the ER membrane; the majority of the la rge subunits remained membrane-bound. On the basis of these results, we pro pose that large ribosomal subunit release from the ER membrane is regulated independently of protein translocation.