Unpaired structures in SCA10 (ATTCT)n·(AGAAT)n repeats

Vladimir N. Potaman, John Bissler, Vera I. Hashem, Elena A. Oussatcheva, Lu Lu, Luda S. Shlyakhtenko, Yuri L. Lyubchenko, Tohru Matsuura, Tetsuo Ashizawa, Michael Leffak, Craig J. Benham, Richard R. Sinden

Research output: Contribution to journalArticle

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Abstract

A number of human hereditary diseases have been associated with the instability of DNA repeats in the genome. Recently, spinocerebellar ataxia type 10 has been associated with expansion of the pentanucleotide repeat (ATTCT)n·(AGAAT)n from a normal range of ten to 22 to as many as 4500 copies. The structural properties of this repeat cloned in circular plasmids were studied by a variety of methods. Two-dimensional gel electrophoresis and atomic force microscopy detected local DNA unpairing in supercoiled plasmids. Chemical probing analysis indicated that, at moderate superhelical densities, the (ATTCT)n·(AGAAT)n repeat forms an unpaired region, which further extends into adjacent A+T-rich flanking sequences at higher superhelical densities. The superhelical energy required to initiate duplex unpairing is essentially length-independent from eight to 46 repeats. In plasmids containing five repeats, minimal unpairing of (ATTCT)5·(AGAAT)5 occurred while 2D gel analysis and chemical probing indicate greater unpairing in A+T-rich sequences in other regions of the plasmid. The observed experimental results are consistent with a statistical mechanical, computational analysis of these supercoiled plasmids. For plasmids containing 29 repeats, which is just above the normal human size range, flanked by an A+T-rich sequence, atomic force microscopy detected the formation of a locally condensed structure at high superhelical densities. However, even at high superhelical densities, DNA strands within the presumably compact A+T-rich region were accessible to small chemicals and oligonucleotide hybridization. Thus, DNA strands in this "collapsed structure" remain unpaired and accessible for interaction with other molecules. The unpaired DNA structure functioned as an aberrant replication origin, in that it supported complete plasmid replication in a HeLa cell extract. A model is proposed in which unscheduled or aberrant DNA replication is a critical step in the expansion mutation.

Original languageEnglish (US)
Pages (from-to)1095-1111
Number of pages17
JournalJournal of Molecular Biology
Volume326
Issue number4
DOIs
StatePublished - Feb 28 2003
Externally publishedYes

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Plasmids
Atomic Force Microscopy
DNA
Superhelical DNA
Inborn Genetic Diseases
Replication Origin
Electrophoresis, Gel, Two-Dimensional
Cell Extracts
DNA Replication
HeLa Cells
Oligonucleotides
Microsatellite Repeats
Reference Values
Gels
Genome
Mutation

All Science Journal Classification (ASJC) codes

  • Structural Biology
  • Molecular Biology

Cite this

Potaman, V. N., Bissler, J., Hashem, V. I., Oussatcheva, E. A., Lu, L., Shlyakhtenko, L. S., ... Sinden, R. R. (2003). Unpaired structures in SCA10 (ATTCT)n·(AGAAT)n repeats. Journal of Molecular Biology, 326(4), 1095-1111. https://doi.org/10.1016/S0022-2836(03)00037-8

Unpaired structures in SCA10 (ATTCT)n·(AGAAT)n repeats. / Potaman, Vladimir N.; Bissler, John; Hashem, Vera I.; Oussatcheva, Elena A.; Lu, Lu; Shlyakhtenko, Luda S.; Lyubchenko, Yuri L.; Matsuura, Tohru; Ashizawa, Tetsuo; Leffak, Michael; Benham, Craig J.; Sinden, Richard R.

In: Journal of Molecular Biology, Vol. 326, No. 4, 28.02.2003, p. 1095-1111.

Research output: Contribution to journalArticle

Potaman, VN, Bissler, J, Hashem, VI, Oussatcheva, EA, Lu, L, Shlyakhtenko, LS, Lyubchenko, YL, Matsuura, T, Ashizawa, T, Leffak, M, Benham, CJ & Sinden, RR 2003, 'Unpaired structures in SCA10 (ATTCT)n·(AGAAT)n repeats', Journal of Molecular Biology, vol. 326, no. 4, pp. 1095-1111. https://doi.org/10.1016/S0022-2836(03)00037-8
Potaman VN, Bissler J, Hashem VI, Oussatcheva EA, Lu L, Shlyakhtenko LS et al. Unpaired structures in SCA10 (ATTCT)n·(AGAAT)n repeats. Journal of Molecular Biology. 2003 Feb 28;326(4):1095-1111. https://doi.org/10.1016/S0022-2836(03)00037-8
Potaman, Vladimir N. ; Bissler, John ; Hashem, Vera I. ; Oussatcheva, Elena A. ; Lu, Lu ; Shlyakhtenko, Luda S. ; Lyubchenko, Yuri L. ; Matsuura, Tohru ; Ashizawa, Tetsuo ; Leffak, Michael ; Benham, Craig J. ; Sinden, Richard R. / Unpaired structures in SCA10 (ATTCT)n·(AGAAT)n repeats. In: Journal of Molecular Biology. 2003 ; Vol. 326, No. 4. pp. 1095-1111.
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