Extrachromosomal double minutes (DM) bear amplified genes that contribute to the malignancy of human cancer cells. A novel intracellular behavior of DMs resulted in their selective entrapment within micronuclei; opening the vista, this could perturb the cancer cell phenotype if genes located on DMs were expressed in micronuclei. Here, using fluorescence in situ hybridization, we detected transcripts in DM-enriched micronuclei. Visualization of DMs and their transcripts in live cells showed that DMs are as actively transcribed in the micronuclei and nuclei. Moreover, pulse-incorporated bromouridine was detected in the micronuclei, and the transcripts eventually exited from the micronuclei, similar to the behavior of nuclear transcripts. This apparently normal pattern of gene expression in DM-enriched micronuclei was restricted to micronuclei associated with lamin B, and lamin B association was more frequent for micronuclei that incorporated DMs than for those that incorporated a chromosome arm. The frequency of lamin B -associated micronuclei increased after entry into S phase, and accordingly, there was a concomitant increase in transcription in micronuclei. Taken together, these results indicate that the expression of genes on DMs can be temporally altered by their incorporation into micronuclei. This may be relevant for a broad spectrum of other extrachromosomal elements. (Mol Cancer Res 2007;5(7):695 -704)

We used fluorescence in situ hybridization (FISH) to detect c-myc RNA subspecies in human COLO 320DM tumor cells. Although the FISH procedure removed the majority of RNAs from the nucleolus, c-myc RNA continued to be detected in both the nucleoplasm and nucleolus. This finding suggests stable association between c-myc RNA and the nucleolus. Nucleolar accumulation of c-myc RNA appeared to be temporally regulated by cell-cycle progression. Hybridization with exon- and strand-specific RNA probes indicated that the non-protein coding exon 1 plays a novel role in determining the subnuclear localization of c-myc RNA. Antisense RNA targeting exon 2 localized only with nucleoplasmic foci, where it might interact with the sense strand. Thus, c-myc gene expression may be regulated by intranuclear localization of its RNA.