Detection and cDNA cloning of H-strand mitochondrial regulatory region RNAs in cultured human cells and human tissues.
In the mitochondrion, essential genetic elements for replication and transcription are mostly housed within a shortsegment of its DNA located between tRNA(Phe) and tRNA(Pro) genes, which is called mitochondrial regulatoryregion (mrr). RNAs are known to be transcribed from mrr, thestructures and the functions of which are yet to be fullycharacterized.We detected ca. 1.3 kb H-strand transcripts of mrr (mrrH-RNAs),and 0.2 kb L-strand transcripts of mrr (mrrL-RNAs) in varioushuman cultured cells and tissues using double stranded mrrDNAprobes. The steady state levels of mrrL-RNAs were generally highin cultured cells, while they varied among tissues. On the otherhand, the levels of mrrH-RNAs varied among tissues and amongcultured cells. A tendency was observed in these cells andtissues that a high level of mrrL-RNA is associated with cellproliferation, and a high level of mrrH-RNA withdifferentiation. Several cDNA clones to 1.3 kb mrrH-RNA were obtained from humanskeletal muscle polyadenylated RNAs. The 5' terminus of the 1.3 kb RNA was determined to be at nucleotide position 15953 whichis immediately downstream of tRNA(Thr) sequence.Polyadenylation site for most of the clones was demonstrated tobe at nucleotide position 576 which is immediately upstream oftRNA(Phe) sequence. The longest cDNA insert obtained was 1177 bps long spanning from nucleotide positions 15969 to 576 which could code for a peptide of 76 amino acids. The cDNAs isolatedhere are the first cDNA clones reported to human mrrH-RNAs.These results, together with previous results, furthersubstantiate that polyadenylated mrrH- and mrrL-RNAs are commonly present at varying levels among human tissues andcells. The 3' end sequences of the cloned mrrH-cDNA provideswith insights into the mechanisms of transcription termination.The cDNA clones will provide tools to further the study of thefunction of mrr RNAs.
Nakamichi
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Cell Technology Center, Meiji Institute of Health Sciences and RITE Meinyu Branch Laboratory, Meiji Milk Products Co., Odawara, Japan, 250-0862.
Ito
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Maeda
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Matsumura
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