Skip to main content
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1992 Jun;12(6):2561–2569. doi: 10.1128/mcb.12.6.2561

Saccharomyces cerevisiae contains an RNase MRP that cleaves at a conserved mitochondrial RNA sequence implicated in replication priming.

L L Stohl 1, D A Clayton 1
PMCID: PMC364449  PMID: 1588958

Abstract

Yeast mitochondrial DNA contains multiple promoters that sponsor different levels of transcription. Several promoters are individually located immediately adjacent to presumed origins of replication and have been suggested to play a role in priming of DNA replication. Although yeast mitochondrial DNA replication origins have not been extensively characterized at the primary sequence level, a common feature of these putative origins is the occurrence of a short guanosine-rich region in the priming strand downstream of the transcriptional start site. This situation is reminiscent of vertebrate mitochondrial DNA origins and raises the possibility of common features of origin function. In the case of human and mouse cells, there exists an RNA processing activity with the capacity to cleave at a guanosine-rich mitochondrial RNA sequence at an origin; we therefore sought the existence of a yeast endoribonuclease that had such a specificity. Whole cell and mitochondrial extracts of Saccharomyces cerevisiae contain an RNase that cleaves yeast mitochondrial RNA in a site-specific manner similar to that of the human and mouse RNA processing activity RNase MRP. The exact location of cleavage within yeast mitochondrial RNA corresponds to a mapped site of transition from RNA to DNA synthesis. The yeast activity also cleaved mammalian mitochondrial RNA in a fashion similar to that of the mammalian RNase MRPs. The yeast endonuclease is a ribonucleoprotein, as judged by its sensitivity to nucleases and proteinase, and it was present in yeast strains lacking mitochondrial DNA, which demonstrated that all components required for in vitro cleavage are encoded by nuclear genes. We conclude that this RNase is the yeast RNase MRP.

Full text

PDF
2561

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Baldacci G., Chérif-Zahar B., Bernardi G. The initiation of DNA replication in the mitochondrial genome of yeast. EMBO J. 1984 Sep;3(9):2115–2120. doi: 10.1002/j.1460-2075.1984.tb02099.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bennett J. L., Clayton D. A. Efficient site-specific cleavage by RNase MRP requires interaction with two evolutionarily conserved mitochondrial RNA sequences. Mol Cell Biol. 1990 May;10(5):2191–2201. doi: 10.1128/mcb.10.5.2191. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Chang D. D., Clayton D. A. A mammalian mitochondrial RNA processing activity contains nucleus-encoded RNA. Science. 1987 Mar 6;235(4793):1178–1184. doi: 10.1126/science.2434997. [DOI] [PubMed] [Google Scholar]
  4. Chang D. D., Clayton D. A. A novel endoribonuclease cleaves at a priming site of mouse mitochondrial DNA replication. EMBO J. 1987 Feb;6(2):409–417. doi: 10.1002/j.1460-2075.1987.tb04770.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chang D. D., Clayton D. A. Mouse RNAase MRP RNA is encoded by a nuclear gene and contains a decamer sequence complementary to a conserved region of mitochondrial RNA substrate. Cell. 1989 Jan 13;56(1):131–139. doi: 10.1016/0092-8674(89)90991-4. [DOI] [PubMed] [Google Scholar]
  6. Chang D. D., Clayton D. A. Priming of human mitochondrial DNA replication occurs at the light-strand promoter. Proc Natl Acad Sci U S A. 1985 Jan;82(2):351–355. doi: 10.1073/pnas.82.2.351. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chang D. D., Hauswirth W. W., Clayton D. A. Replication priming and transcription initiate from precisely the same site in mouse mitochondrial DNA. EMBO J. 1985 Jun;4(6):1559–1567. doi: 10.1002/j.1460-2075.1985.tb03817.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Clayton D. A. Replication of animal mitochondrial DNA. Cell. 1982 Apr;28(4):693–705. doi: 10.1016/0092-8674(82)90049-6. [DOI] [PubMed] [Google Scholar]
  9. England T. E., Uhlenbeck O. C. 3'-terminal labelling of RNA with T4 RNA ligase. Nature. 1978 Oct 12;275(5680):560–561. doi: 10.1038/275560a0. [DOI] [PubMed] [Google Scholar]
  10. Fangman W. L., Dujon B. Yeast mitochondrial genomes consisting of only A.T base pairs replicate and exhibit suppressiveness. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7156–7160. doi: 10.1073/pnas.81.22.7156. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Faugeron-Fonty G., Le Van Kim C., de Zamaroczy M., Goursot R., Bernardi G. A comparative study of the ori sequences from the mitochondrial genomes of twenty wild-type yeast strains. Gene. 1984 Dec;32(3):459–473. doi: 10.1016/0378-1119(84)90020-9. [DOI] [PubMed] [Google Scholar]
  12. Gold H. A., Topper J. N., Clayton D. A., Craft J. The RNA processing enzyme RNase MRP is identical to the Th RNP and related to RNase P. Science. 1989 Sep 22;245(4924):1377–1380. doi: 10.1126/science.2476849. [DOI] [PubMed] [Google Scholar]
  13. Greenleaf A. L., Kelly J. L., Lehman I. R. Yeast RPO41 gene product is required for transcription and maintenance of the mitochondrial genome. Proc Natl Acad Sci U S A. 1986 May;83(10):3391–3394. doi: 10.1073/pnas.83.10.3391. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hollingsworth M. J., Martin N. C. RNase P activity in the mitochondria of Saccharomyces cerevisiae depends on both mitochondrion and nucleus-encoded components. Mol Cell Biol. 1986 Apr;6(4):1058–1064. doi: 10.1128/mcb.6.4.1058. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Karwan R., Bennett J. L., Clayton D. A. Nuclear RNase MRP processes RNA at multiple discrete sites: interaction with an upstream G box is required for subsequent downstream cleavages. Genes Dev. 1991 Jul;5(7):1264–1276. doi: 10.1101/gad.5.7.1264. [DOI] [PubMed] [Google Scholar]
  16. Lambowitz A. M. Preparation and analysis of mitochondrial ribosomes. Methods Enzymol. 1979;59:421–433. doi: 10.1016/0076-6879(79)59103-4. [DOI] [PubMed] [Google Scholar]
  17. Lee J. Y., Engelke D. R. Partial characterization of an RNA component that copurifies with Saccharomyces cerevisiae RNase P. Mol Cell Biol. 1989 Jun;9(6):2536–2543. doi: 10.1128/mcb.9.6.2536. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Lin R. J., Newman A. J., Cheng S. C., Abelson J. Yeast mRNA splicing in vitro. J Biol Chem. 1985 Nov 25;260(27):14780–14792. [PubMed] [Google Scholar]
  19. Miller D. L., Martin N. C. Characterization of the yeast mitochondrial locus necessary for tRNA biosynthesis: DNA sequence analysis and identification of a new transcript. Cell. 1983 Oct;34(3):911–917. doi: 10.1016/0092-8674(83)90548-2. [DOI] [PubMed] [Google Scholar]
  20. Morales M. J., Wise C. A., Hollingsworth M. J., Martin N. C. Characterization of yeast mitochondrial RNase P: an intact RNA subunit is not essential for activity in vitro. Nucleic Acids Res. 1989 Sep 12;17(17):6865–6881. doi: 10.1093/nar/17.17.6865. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Parikh V. S., Morgan M. M., Scott R., Clements L. S., Butow R. A. The mitochondrial genotype can influence nuclear gene expression in yeast. Science. 1987 Jan 30;235(4788):576–580. doi: 10.1126/science.3027892. [DOI] [PubMed] [Google Scholar]
  22. Read S. M., Northcote D. H. Minimization of variation in the response to different proteins of the Coomassie blue G dye-binding assay for protein. Anal Biochem. 1981 Sep 1;116(1):53–64. doi: 10.1016/0003-2697(81)90321-3. [DOI] [PubMed] [Google Scholar]
  23. Schinkel A. H., Tabak H. F. Mitochondrial RNA polymerase: dual role in transcription and replication. Trends Genet. 1989 May;5(5):149–154. doi: 10.1016/0168-9525(89)90056-5. [DOI] [PubMed] [Google Scholar]
  24. Topper J. N., Clayton D. A. Characterization of human MRP/Th RNA and its nuclear gene: full length MRP/Th RNA is an active endoribonuclease when assembled as an RNP. Nucleic Acids Res. 1990 Feb 25;18(4):793–799. doi: 10.1093/nar/18.4.793. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Wong T. W., Clayton D. A. DNA primase of human mitochondria is associated with structural RNA that is essential for enzymatic activity. Cell. 1986 Jun 20;45(6):817–825. doi: 10.1016/0092-8674(86)90556-8. [DOI] [PubMed] [Google Scholar]
  26. Wong T. W., Clayton D. A. In vitro replication of human mitochondrial DNA: accurate initiation at the origin of light-strand synthesis. Cell. 1985 Oct;42(3):951–958. doi: 10.1016/0092-8674(85)90291-0. [DOI] [PubMed] [Google Scholar]
  27. de Zamaroczy M., Faugeron-Fonty G., Baldacci G., Goursot R., Bernardi G. The ori sequences of the mitochondrial genome of a wild-type yeast strain: number, location, orientation and structure. Gene. 1984 Dec;32(3):439–457. doi: 10.1016/0378-1119(84)90019-2. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES