Conformational Variants of Duplex DNA Correlated with Cytosine-rich Chromosomal Fragile Sites

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

doi:10.1074/jbc.m806866200

Cited by 44 publications

(67 citation statements)

References 33 publications

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“…But the methylase probing studies indicate that the fragile zones are not single stranded for a large fraction of the time. Transient non-B conformations (called B/A-intermediate DNA) at the CCND1-MTC region and BCL2-MBR have been documented elsewhere, though these are still within the range of duplex conformations (4,9).…”

Section: Discussionmentioning

confidence: 99%

“…1A and 2A). These zones are 100-to 1,000-fold more sensitive to breakage than the adjacent DNA regions located, for example, only 20 bp to either side of the fragile zone (2)(3)(4)(5)(6)(7). Hence, the DNA sequences surrounding each oncogene are not equally sensitive to breakage.…”

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confidence: 99%

“…Namely, why are the CG sites in these fragile zones more sensitive to breakage than the CG sites in DNA only 20 to 100 bp away? The presence of altered DNA structure within the zones represents one possibility, though the in vivo longevity of such altered DNA structures is unclear (4,9).…”

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confidence: 99%

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Human Lymphoid Translocation Fragile Zones Are Hypomethylated and Have Accessible Chromatin

Lieber

Tsai

et al. 2015

Molecular and Cellular Biology

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Chromosomal translocations are a hallmark of hematopoietic malignancies. CG motifs within translocation fragile zones (typically 20 to 600 bp in size) are prone to chromosomal translocation in lymphomas. Here we demonstrate that the CG motifs in human translocation fragile zones are hypomethylated relative to the adjacent DNA. Using a methyltransferase footprinting assay on isolated nuclei (in vitro), we find that the chromatin at these fragile zones is accessible. We also examined in vivo accessibility using cellular expression of a prokaryotic methylase. Based on this assay, which measures accessibility over a much longer time interval than is possible with in vitro methods, these fragile zones were found to be more accessible than the adjacent DNA. Because DNA within the fragile zones can be methylated by both cellular and exogenous methyltransferases, the fragile zones are predominantly in a duplex DNA conformation. These observations permit more-refined models for why these zones are 100-to 1,000-fold more prone to undergo chromosomal translocation than the adjacent regions.C hromosomal translocations are seminal events in the development of many hematologic malignancies (1). Many hematopoietic malignancies are associated with recurrent reciprocal translocations and their resulting gene fusions or amplifications, which guide diagnosis, assessment of prognosis, and treatment. One example is the BCL2-IGH translocation, t(14;18)(q32;q21), observed in 85% of follicular lymphomas (FL) and 30% of diffuse large B cell lymphomas (DLBCL). A second example is the CCND1-IGH translocation, t(11;14)(q13;q32), observed in almost all mantle cell lymphomas (MCL). Oncogenes near sites of recurrent translocation often confer a selective growth advantage (2). For unknown reasons, human chromosomal translocation breakpoints near these oncogenes frequently cluster within tight DNA zones (often only 20 to 600 bp in length) known as fragile zones (see the green starbursts in the upper portion of Fig. 1A and 2A). These zones are 100-to 1,000-fold more sensitive to breakage than the adjacent DNA regions located, for example, only 20 bp to either side of the fragile zone (2-7). Hence, the DNA sequences surrounding each oncogene are not equally sensitive to breakage. The molecular basis for such an extreme breakage propensity within such a short zone of DNA is unknown, despite our progress on elucidation of other aspects of the mechanism described below (8).We have previously reported that breakpoints of human lymphoid translocation fragile zones are not randomly distributed (Fig. 1A and 2A). Pro-B/pre-B cell-stage translocations show a breakage propensity for the CG (commonly called CpG but designated CG in this paper) DNA sequence motif within the 20 to 600-bp fragile zones (3). We have proposed a model in which activation-induced deaminase (AID) initiates double-strand breaks (DSBs) by acting at the methylated form of these CG sites in single-stranded DNA (ssDNA) to deaminate the 5-methylcytosine to yield a T-G mismatch (3). The r...

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Human Lymphoid Translocation Fragile Zones Are Hypomethylated and Have Accessible Chromatin

Lieber

Tsai

et al. 2015

Molecular and Cellular Biology

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“…Top1 reduction is likely to retard relaxation of negative supercoil. Excessive negative supercoil of DNA facilitates local structural alterations including R-loop, stem-loop, cruciform, triplex, and other non-B DNA structures in DNA rich in palindromes or G/C stretches like the S region, which probably inhibits the religation step of Top1 after cleavage (3,(8)(9)(10) (Figs. S7 and S8).…”

Section: Discussionmentioning

confidence: 99%

“…Lieber and colleagues reported that the bisulfite treatment of nuclear DNA from switch-induced spleen cells converted dC to dU in the S region, indicating the presence of single stranded DNA in the S region in activated B cells (6,7). R-loop may not survive long but lead to the formation of non-B structures including stem-loop, cruciform, and triplex in the S region because of abundant inverted repeats (3,(8)(9)(10).…”

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AID-induced decrease in topoisomerase 1 induces DNA structural alteration and DNA cleavage for class switch recombination

Kobayashi

Aida

Nagaoka

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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To initiate class switch recombination (CSR) activation-induced cytidine deaminase (AID) induces staggered nick cleavage in the S region, which lies 5 to each Ig constant region gene and is rich in palindromic sequences. Topoisomerase 1 (Top1) controls the supercoiling of DNA by nicking, rotating, and religating one strand of DNA. Curiously, Top1 reduction or AID overexpression causes the genomic instability. Here, we report that the inactivation of Top1 by its specific inhibitor camptothecin drastically blocked both the S region cleavage and CSR, indicating that Top1 is responsible for the S region cleavage in CSR. Surprisingly, AID expression suppressed Top1 mRNA translation and reduced its protein level. In addition, the decrease in the Top1 protein by RNA-mediated knockdown augmented the AID-dependent S region cleavage, as well as CSR. Furthermore, Top1 reduction altered DNA structure of the S region. Taken together, AID-induced Top1 reduction alters S region DNA structure probably to non-B form, on which Top1 can introduce nicks but cannot religate, resulting in S region cleavage.camptothecin ͉ non-B DNA ͉ translation suppression

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Nonhomologous DNA End Joining (NHEJ) and Chromosomal Translocations in Humans

Lieber

et al. 2009

Subcellular Biochemistry

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Estimates are that double-strand breaks (DSBs) arise in dividing cells about ten times per cell per day. Causes include replication across a nick, free radicals of oxidative metabolism, ionizing radiation, and inadvertent action by enzymes of DNA metabolism (such as failures of type II topoisomerases or cleavage by recombinases at pseudo sequences that look sufficiently similar to the physiologic ones). There are two major double-strand break repair pathways. Homologous recombination (HR) can repair double-strand breaks, but only during S phase and only if there is sufficient homology, usually more than 100 bp. The more general and commonly used pathway is nonhomologous DNA end joining, abbreviated NHEJ. NHEJ can repair a DSB at any time during the cell cycle and does not require any homology, though a few nucleotides of terminal microhomology are often utilized by the NHEJ enzymes, if present. The enzymes of NHEJ include Ku, DNA-PKcs, Artemis, polymerase mu, polymerse lambda, XLF (also called Cernunnos), XRCC4, and DNA ligase IV. These enzymes constitute what some call the classical NHEJ pathway, and in wild type cells, the vast majority of joining events appear to proceed using these components. In rare mammalian cell mutants of ligase IV, for example, much less efficient joining occurs using one of the remaining two ligases, ligase I or III, in a manner that is much more reliant on longer terminal microhomology lengths, such as 7 to 15 nt; this is usually called alternative NHEJ, back-up NHEJ, or microhomology-mediated end joining (MMEJ). NHEJ is present in many prokaryotes, as well as all as eukaryotes, and very similar mechanistic flexibility evolved both convergently and divergently. When two double-strand breaks occur on different chromosomes, then the rejoining is almost always done by NHEJ. The causes of the DSBs in lymphomas most often involve the RAG or AID enzymes that function in the specialized processes of antigen receptor gene rearrangement.

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Conformational Variants of Duplex DNA Correlated with Cytosine-rich Chromosomal Fragile Sites

Cited by 44 publications

References 33 publications

Human Lymphoid Translocation Fragile Zones Are Hypomethylated and Have Accessible Chromatin

Human Lymphoid Translocation Fragile Zones Are Hypomethylated and Have Accessible Chromatin

AID-induced decrease in topoisomerase 1 induces DNA structural alteration and DNA cleavage for class switch recombination

Nonhomologous DNA End Joining (NHEJ) and Chromosomal Translocations in Humans

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