Conformational variants of duplex DNA correlated with cytosine-rich chromosomal fragile sites

Albert G. Tsai, Aaron E. Engelhart, Ma'mon M. Hatmal, Sabrina I. Houston, Nicholas V. Hud, Ian S. Haworth, Michael R. Lieber

Research output: Contribution to journalArticle

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Abstract

We found that several major chromosomal fragile sites in human lymphomas, including the bcl-2 major breakpoint region, bcl-1 major translocation cluster, and c-Myc exon 1-intron 1 boundary, contain distinctive sequences of consecutive cytosines exhibiting a high degree of reactivity with the structure-specific chemical probe bisulfite. To assess the inherent structural variability of duplex DNA in these regions and to determine the range of structures reactive to bisulfite, we have performed bisulfite probing on genomic DNA in vitro and in situ; on duplex DNA in supercoiled and linearized plasmids; and on oligonucleotide DNA/DNA and DNA/2′-O-methyl RNA duplexes. Bisulfite is significantly more reactive at the frayed ends of DNA duplexes, which is expected given that bisulfite is an established probe of single-stranded DNA. We observed that bisulfite also distinguishes between more subtle sequence/structural differences in duplex DNA. Supercoiled plasmids are more reactive than linear DNA; and sequences containing consecutive cytosines, namely GGGCCC, are more reactive than those with alternating guanine and cytosine, namely GCGCGC. Circular dichroism and x-ray crystallography show that the GGGCCC sequence forms an intermediate B/A structure. Molecular dynamics simulations also predict an intermediate B/A structure-for this sequence, and probe calculations suggest greater bisulfite accessibility of cytosine bases in the intermediate B/A structure over canonical B- or A-form DNA. Electrostatic calculations reveal that consecutive cytosine bases create electropositive patches in the major groove, predicting enhanced localization of the bisulfite anion at homo-C tracts over alternating G/ C sequences. These characteristics of homo-C tracts in duplex DNA may be associated with DNA-protein interactions in vivo that predispose certain genomic regions to chromosomal fragility.

Original languageEnglish (US)
Pages (from-to)7157-7164
Number of pages8
JournalJournal of Biological Chemistry
Volume284
Issue number11
DOIs
StatePublished - Mar 13 2009

Fingerprint

Cytosine
DNA
Chromosome Fragility
Plasmids
A-Form DNA
Superhelical DNA
Crystallography
hydrogen sulfite
B-Form DNA
Single-Stranded DNA
Guanine
Molecular Dynamics Simulation
Circular Dichroism
Static Electricity
Oligonucleotides
Introns
Anions
Exons
Lymphoma
X-Rays

Cite this

Tsai, A. G., Engelhart, A. E., Hatmal, M. M., Houston, S. I., Hud, N. V., Haworth, I. S., & Lieber, M. R. (2009). Conformational variants of duplex DNA correlated with cytosine-rich chromosomal fragile sites. Journal of Biological Chemistry, 284(11), 7157-7164. https://doi.org/10.1074/jbc.M806866200

Conformational variants of duplex DNA correlated with cytosine-rich chromosomal fragile sites. / Tsai, Albert G.; Engelhart, Aaron E.; Hatmal, Ma'mon M.; Houston, Sabrina I.; Hud, Nicholas V.; Haworth, Ian S.; Lieber, Michael R.

In: Journal of Biological Chemistry, Vol. 284, No. 11, 13.03.2009, p. 7157-7164.

Research output: Contribution to journalArticle

Tsai, Albert G. ; Engelhart, Aaron E. ; Hatmal, Ma'mon M. ; Houston, Sabrina I. ; Hud, Nicholas V. ; Haworth, Ian S. ; Lieber, Michael R. / Conformational variants of duplex DNA correlated with cytosine-rich chromosomal fragile sites. In: Journal of Biological Chemistry. 2009 ; Vol. 284, No. 11. pp. 7157-7164.
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AB - We found that several major chromosomal fragile sites in human lymphomas, including the bcl-2 major breakpoint region, bcl-1 major translocation cluster, and c-Myc exon 1-intron 1 boundary, contain distinctive sequences of consecutive cytosines exhibiting a high degree of reactivity with the structure-specific chemical probe bisulfite. To assess the inherent structural variability of duplex DNA in these regions and to determine the range of structures reactive to bisulfite, we have performed bisulfite probing on genomic DNA in vitro and in situ; on duplex DNA in supercoiled and linearized plasmids; and on oligonucleotide DNA/DNA and DNA/2′-O-methyl RNA duplexes. Bisulfite is significantly more reactive at the frayed ends of DNA duplexes, which is expected given that bisulfite is an established probe of single-stranded DNA. We observed that bisulfite also distinguishes between more subtle sequence/structural differences in duplex DNA. Supercoiled plasmids are more reactive than linear DNA; and sequences containing consecutive cytosines, namely GGGCCC, are more reactive than those with alternating guanine and cytosine, namely GCGCGC. Circular dichroism and x-ray crystallography show that the GGGCCC sequence forms an intermediate B/A structure. Molecular dynamics simulations also predict an intermediate B/A structure-for this sequence, and probe calculations suggest greater bisulfite accessibility of cytosine bases in the intermediate B/A structure over canonical B- or A-form DNA. Electrostatic calculations reveal that consecutive cytosine bases create electropositive patches in the major groove, predicting enhanced localization of the bisulfite anion at homo-C tracts over alternating G/ C sequences. These characteristics of homo-C tracts in duplex DNA may be associated with DNA-protein interactions in vivo that predispose certain genomic regions to chromosomal fragility.

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