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Review
, 5 (1), e1000327

A Microhomology-Mediated Break-Induced Replication Model for the Origin of Human Copy Number Variation

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Review

A Microhomology-Mediated Break-Induced Replication Model for the Origin of Human Copy Number Variation

P J Hastings et al. PLoS Genet.

Abstract

Chromosome structural changes with nonrecurrent endpoints associated with genomic disorders offer windows into the mechanism of origin of copy number variation (CNV). A recent report of nonrecurrent duplications associated with Pelizaeus-Merzbacher disease identified three distinctive characteristics. First, the majority of events can be seen to be complex, showing discontinuous duplications mixed with deletions, inverted duplications, and triplications. Second, junctions at endpoints show microhomology of 2-5 base pairs (bp). Third, endpoints occur near pre-existing low copy repeats (LCRs). Using these observations and evidence from DNA repair in other organisms, we derive a model of microhomology-mediated break-induced replication (MMBIR) for the origin of CNV and, ultimately, of LCRs. We propose that breakage of replication forks in stressed cells that are deficient in homologous recombination induces an aberrant repair process with features of break-induced replication (BIR). Under these circumstances, single-strand 3' tails from broken replication forks will anneal with microhomology on any single-stranded DNA nearby, priming low-processivity polymerization with multiple template switches generating complex rearrangements, and eventual re-establishment of processive replication.

Figures

Figure 1
Figure 1. In silico analyses revealed complex genomic architecture in regions of nonrecurrent rearrangement.
(A) The ∼3 Mb surrounding the PLP1 gene and (B) the ∼4 Mb surrounding the MECP2 gene on the X chromosome contain numerous LCRs in various orientations ,. LCRs are represented by the colored block arrows, and like LCR copies are designated by color and letter for a given sequence. Orientation is depicted by the direction of the block arrow.
Figure 2
Figure 2. Complex rearrangements involving PLP1 detected by junction analysis (A) and oligonucleotide array comparative genomic hybridization analysis (B) .
(A) A complex duplication of the PLP1 region detected by outward facing polymerase chain reaction. Panel (i) shows the PLP1 region with the positions of the outward facing primers. The structure of the duplicated region is shown in (ii), with an enlargement of the complex junction region in (iii). Two or three bp of microhomology, shown by the letters A, C, G and T, were found at the breakpoint junctions (open arrows). (B) Deletion and duplications found in two patients with Pelizaeus-Merzbacher disease and their carrier mother , shown by comparative genomic hybridization. A ∼190-kb deletion is followed by a ∼9-kb segment with no copy-number change, and an interrupted ∼190-kb duplication was detected (i). Panel (ii) shows enlargement of the array revealing interruption of the ∼190-kb duplication. In each horizontal yellow box above, blue lines represent an average of the data points. Red data points indicate copy-number gains, green data points indicate losses, and black data points indicate no copy-number change. The y-axes show relative hybridization; genomic position is on the x-axis. Panel (iii) summarizes the structure based on comparative genomic hybridization where a green box shows the region deleted, red boxes show the regions duplicated, and black lines show regions of no change.
Figure 3
Figure 3. Complex genomic rearrangements at PLP1 seen in patients with Pelizaeus-Merzbacher disease, illustrating long-range as well as short-range complexity.
Duplications are shown in red, deletions in green, triplications in blue, and no copy number change in black. The figure is not drawn to scale. Approximate positions are given relative to PLP1.
Figure 4
Figure 4. Repair of a collapsed replication fork by BIR.
When a replication fork encounters a nick in a template strand (A) (arrowhead), one arm of the fork breaks off (red), producing a collapsed fork (B). At the single double-strand end, the 5′ strand is resected, giving a 3′ overhang (C). The 3′ single-strand end invades the sister molecule (blue), forming a D-loop (D), which subsequently becomes a replication fork with both leading and lagging strand replication (E). There is a Holliday junction at the site of the D-loop. Migration of the Holliday junction, or some other helicase activity, separates the extended double-strand end from its templates (F). The separated end is again processed to give a 3′ single-strand end, which again invades the sister, and forms a replication fork (G). Eventually the replication fork becomes fully processive, and continues replication to the chromosome end (H and I). This process is shown here with the Holliday junction following the fork so that newly formed strands are segregated together (conservative segregation) (H). Each line represents a DNA nucleotide chain (strand). Polarity is indicated by half arrows on 3′ end. New DNA synthesis is shown by dashed lines. The publications on which this model is based are cited in the text.
Figure 5
Figure 5. MMBIR.
The figure shows successive switches to different genomic positions (distinguished by color) forming microhomology junctions (arrows). For clarity, the nature of the single-stranded regions of annealing is not defined (see text). (A) shows the broken arm of a collapsed replication fork, which forms a new low-processivity fork as shown at (B). The extended end dissociates repeatedly ((C and E) shown with 5′-ends resected) and reforms the fork on different templates (D and F). In (F), the switch returns to the original sister chromatid (blue), forming a processive replication fork that completes replication. (G) shows the final product containing sequence from different genomic regions. Each line represents a DNA nucleotide chain (strand). Polarity is indicated by half arrows on 3′ end. Whether the return to the sister chromatid occurs in front of or behind the position of the original collapse determines whether there is a deletion or duplication (see Table 2).

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