Localization of the newly initiated and processed ribosomal primary transcripts during the mitotic cycle in Physarum...

We analyzed the fate of the rRNA released as a result of the prophasic disintegration of the nucleolus, during the "closed" mitosis of the naturally synchronous plasmodium of Physarum polycephalum. Using a probe complementary to the mature 19S and 26S rRNA, we previously showed that the nucleolus-derived rRNAs are stable in mitosis, mainly associated with numerous fibrillar nucleolar remnants (Pierron and Puvion-Dutilleul, 1993, Exp. Cell Res. 208, 509-517). However, a significant fraction of these mitotic rRNA precursors were also found in more diffuse nuclear, granular zones. In this paper, we trace pre-rRNA molecules in the early stages of their maturation by using probes derived from the 5' external transcribed spacer (5'-ETS), upstream and downstream of the processing site located 1.7 kb from the transcription initiation site. In agreement with a previous S1-mapping study (Blum et al., 1986, Nucleic Acids Res., 10, 4121-4133), we observed that the rRNA transcripts complementary to the 5'-ETS portion downstream of the processing site are stable in mitosis, and we demonstrate that they are specifically associated with the border of the fibrillar nucleolar remnants. On the other hand, the pre-rRNA chains containing the upstream portion of the 5'-ETS become undetectable in mitosis, marking the cessation of rDNA transcription. Since the reappearance of these nascent pre-rRNA molecules signals the resumption of the transcriptional activity, we determined that a burst of rRNA synthesis is taking place within the prenucleolar bodies, immediately after the separation of the daughter nuclei. Taken together, our results illustrate the persistence of the nucleolar components of Physarum, within nucleolar remnants that, although transcriptionally inactive during mitosis, function as independent entities very early in interphase.