Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1989 Jan;9(1):169–176. doi: 10.1128/mcb.9.1.169

Replication of DNA minicircles in kinetoplasts isolated from Crithidia fasciculata: structure of nascent minicircles.

C Sheline 1, T Melendy 1, D S Ray 1
PMCID: PMC362158  PMID: 2927390

Abstract

We have previously described an isolated kinetoplast system from Crithidia fasciculata capable of ATP-dependent replication of kinetoplast DNA minicircles (L. Birkenmeyer and D.S. Ray, J. Biol. Chem. 261: 2362-2368, 1986). We present here the identification of two new minicircle species observed in short pulse-labeling experiments in this system. The earliest labeled minicircle species (component A) contains both nascent H and L strands and is heterogeneous in sedimentation and electrophoretic migration. Component A has characteristics consistent with a Cairns-type structure in which the L strand is the leading strand and the H strand is the lagging strand. The other new species (component B) has a nascent 2.5-kilobase linear L strand with a single discontinuity that mapped to either of two alternative origins located 180 degrees apart on the minicircle map. Component B could be repaired to a covalently closed form by Escherichia coli polymerase I and T4 ligase but not by T4 polymerase and T4 ligase. Even though component B has a single gap in one strand, it had an electrophoretic mobility on an agarose gel (minus ethidium bromide) similar to that of a supercoiled circle with three supertwists. Treatment of component B with topoisomerase II converted it to a form that comigrated with a nicked open circular form (replicative form II). These results indicate that component B is a knotted topoisomer of a kinetoplast DNA minicircle with a single gap in the L strand.

Full text

PDF
175

Images in this article

171Image
on p.171
171Image
on p.171
172Image
on p.172
172Image
on p.172
173Image
on p.173
174Image
on p.174
175Image
on p.175

Selected References

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

  1. Birkenmeyer L., Ray D. S. Replication of kinetoplast DNA in isolated kinetoplasts from Crithidia fasciculata. Identification of minicircle DNA replication intermediates. J Biol Chem. 1986 Feb 15;261(5):2362–2368. [PubMed] [Google Scholar]
  2. Birkenmeyer L., Sugisaki H., Ray D. S. Structural characterization of site-specific discontinuities associated with replication origins of minicircle DNA from Crithidia fasciculata. J Biol Chem. 1987 Feb 15;262(5):2384–2392. [PubMed] [Google Scholar]
  3. Birkenmeyer L., Sugisaki H., Ray D. S. The majority of minicircle DNA in Crithidia fasciculata strain CF-C1 is of a single class with nearly homogeneous DNA sequence. Nucleic Acids Res. 1985 Oct 11;13(19):7107–7118. doi: 10.1093/nar/13.19.7107. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. 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]
  5. Dean F. B., Cozzarelli N. R. Mechanism of strand passage by Escherichia coli topoisomerase I. The role of the required nick in catenation and knotting of duplex DNA. J Biol Chem. 1985 Apr 25;260(8):4984–4994. [PubMed] [Google Scholar]
  6. Englund P. T. Free minicircles of kinetoplast DNA in Crithidia fasciculata. J Biol Chem. 1979 Jun 10;254(11):4895–4900. [PubMed] [Google Scholar]
  7. Englund P. T., Hajduk S. L., Marini J. C. The molecular biology of trypanosomes. Annu Rev Biochem. 1982;51:695–726. doi: 10.1146/annurev.bi.51.070182.003403. [DOI] [PubMed] [Google Scholar]
  8. Hsieh T. Knotting of the circular duplex DNA by type II DNA topoisomerase from Drosophila melanogaster. J Biol Chem. 1983 Jul 10;258(13):8413–8420. [PubMed] [Google Scholar]
  9. Kitchin P. A., Klein V. A., Englund P. T. Intermediates in the replication of kinetoplast DNA minicircles. J Biol Chem. 1985 Mar 25;260(6):3844–3851. [PubMed] [Google Scholar]
  10. Kitchin P. A., Klein V. A., Fein B. I., Englund P. T. Gapped Minicircles. A novel replication intermediate of kinetoplast DNA. J Biol Chem. 1984 Dec 25;259(24):15532–15539. [PubMed] [Google Scholar]
  11. Liu L. F., Liu C. C., Alberts B. M. Type II DNA topoisomerases: enzymes that can unknot a topologically knotted DNA molecule via a reversible double-strand break. Cell. 1980 Mar;19(3):697–707. doi: 10.1016/s0092-8674(80)80046-8. [DOI] [PubMed] [Google Scholar]
  12. Ntambi J. M., Englund P. T. A gap at a unique location in newly replicated kinetoplast DNA minicircles from Trypanosoma equiperdum. J Biol Chem. 1985 May 10;260(9):5574–5579. [PubMed] [Google Scholar]
  13. Ntambi J. M., Marini J. C., Bangs J. D., Hajduk S. L., Jimenez H. E., Kitchin P. A., Klein V. A., Ryan K. A., Englund P. T. Presence of a bent helix in fragments of kinetoplast DNA minicircles from several trypanosomatid species. Mol Biochem Parasitol. 1984 Jul;12(3):273–286. doi: 10.1016/0166-6851(84)90084-7. [DOI] [PubMed] [Google Scholar]
  14. Ntambi J. M., Shapiro T. A., Ryan K. A., Englund P. T. Ribonucleotides associated with a gap in newly replicated kinetoplast DNA minicircles from Trypanosoma equiperdum. J Biol Chem. 1986 Sep 5;261(25):11890–11895. [PubMed] [Google Scholar]
  15. Ray D. S., Hines J. C., Sugisaki H., Sheline C. kDNA minicircles of the major sequence class of C. fasciculata contain a single region of bent helix widely separated from the two origins of replication. Nucleic Acids Res. 1986 Oct 24;14(20):7953–7965. doi: 10.1093/nar/14.20.7953. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ray D. S. Kinetoplast DNA minicircles: high-copy-number mitochondrial plasmids. Plasmid. 1987 May;17(3):177–190. doi: 10.1016/0147-619x(87)90026-6. [DOI] [PubMed] [Google Scholar]
  17. Ryan K. A., Shapiro T. A., Rauch C. A., Englund P. T. Replication of kinetoplast DNA in trypanosomes. Annu Rev Microbiol. 1988;42:339–358. doi: 10.1146/annurev.mi.42.100188.002011. [DOI] [PubMed] [Google Scholar]
  18. Ryan K. A., Shapiro T. A., Rauch C. A., Griffith J. D., Englund P. T. A knotted free minicircle in kinetoplast DNA. Proc Natl Acad Sci U S A. 1988 Aug;85(16):5844–5848. doi: 10.1073/pnas.85.16.5844. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Simpson L. Kinetoplast DNA in trypanosomid flagellates. Int Rev Cytol. 1986;99:119–179. doi: 10.1016/s0074-7696(08)61426-6. [DOI] [PubMed] [Google Scholar]
  20. Simpson L. The mitochondrial genome of kinetoplastid protozoa: genomic organization, transcription, replication, and evolution. Annu Rev Microbiol. 1987;41:363–382. doi: 10.1146/annurev.mi.41.100187.002051. [DOI] [PubMed] [Google Scholar]
  21. Spengler S. J., Stasiak A., Cozzarelli N. R. The stereostructure of knots and catenanes produced by phage lambda integrative recombination: implications for mechanism and DNA structure. Cell. 1985 Aug;42(1):325–334. doi: 10.1016/s0092-8674(85)80128-8. [DOI] [PubMed] [Google Scholar]
  22. Sugisaki H., Ray D. S. DNA sequence of Crithidia fasciculata kinetoplast minicircles. Mol Biochem Parasitol. 1987 Apr;23(3):253–263. doi: 10.1016/0166-6851(87)90032-6. [DOI] [PubMed] [Google Scholar]

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

RESOURCES