Figure 3.

Experimental validation of the expression of CCA-RNAs.

Total RNAs from the indicated human cultured cell lines were subjected to northern blot using the probes shown in Fig. S3. 5S rRNA and cytoplasmic tRNA^LysCUU were analyzed as controls. Detected (N)CCA-RNAs are indicated by lines. In the detection of ccaR-2 using probe #1, we observed a non-specific band (indicated by a single asterisk) that was derived from a cytoplasmic RNA (Fig. 4) and was not regulated by CCA-adding enzyme (Fig. 5). In contrast, detection of nccaR-1 using both probes #1 and #2 identified potential precursor molecules (indicated by double asterisks) that were present in mitochondria (Fig. 4). Given its size, the clearly observed nccaR-1 band (indicated by a line) may correspond with RNAs that lack nucleotide additions, and nucleotide-added sequences are likely represented by the upper smeared bands.