Discrimination of potential and actual RNA polymerase B activity in isolated nuclei during differentiation ofPhysarum polycephalum

Abstract

Isolated nuclei ofPhysarum contain endogenous RNA polymerase activity. We provide evidence for four different states of RNA polymerase B: 1. free enzyme (85%); 2. weakly bound enzyme (10%) and 3. tightly bound enzyme (0-4%), which can be solubilized from isolated nuclei with 0.5 M and 1.5 M NaCl respectively; 4. "initiated" enzyme. The latter fraction (1-5% of the total RNA polymerase B) is not soluble in salt extractions, does not accept external templates, shows high salt optimum for transcription (0.4 M NaCl) and produces by elongation RNA molecules of mainly 10 S. Treatment of isolated nuclei from differentiating cultures with Triton X-100 increases the proportion of the "initiated" enzyme at the expense of the tightly bound enzyme fraction. This indicates a potential transcription control mechanism which operates at the chromatin level and results in variable proportions of silent and transcribing RNA polymerase B molecules during differentiation of Physarum.