Figure 3.

Subcellular localization of RPS2 by in vitro translation/translocation. (A) RPS2 appears to be cytoplasmic. RPS2 RNA (lanes 2–6) and β-lactamase RNA (a positive control for translocation; lanes 7–11) were translated either in the presence (lanes 3–6 and 8–11) or absence (lanes 2 and 7) of dog pancreatic microsomes. Lane 1 shows a no RNA control for translation. The reactions were treated with proteinase K (lanes 4 and 9) or fractionated by ultracentrifugation into precipitate (lanes 5 and 10) and supernatant (lanes 6 and 11) fractions after Na₂CO₃ treatment. The positions of molecular weight markers, RPS2, and β-lactamase are indicated on the left. (B) RPS2 detected in the precipitate fraction is an artifact. The microsomes were either included in the translation reaction as in the standard procedure (cotranslation; lanes 3 and 4) or added after the translation reaction was terminated with cycloheximide (posttranslation; lanes 1 and 2). The reactions were fractionated into precipitate (lanes 1 and 3) and supernatant fractions (lanes 2 and 4) after Na₂CO₃ treatment.