Insertional translocation detected using FISH confirmation of array-comparative genomic hybridization (aCGH) results

Abstract

Insertional translocations (ITs) are rare events that require at least three breaks in the chromosomes involved and thus qualify as complex chromosomal rearrangements (CCR). In the current study, we identified 40 ITs from approximately 18,000 clinical cases (1:500) using array-comparative genomic hybridization (aCGH) in conjunction with fluorescence in situ hybridization (FISH) confirmation of the aCGH findings, and parental follow-up studies. Both submicroscopic and microscopically visible IT events were detected. They were divided into three major categories: (1) simple intrachromosomal and interchromosomal IT resulting in pure segmental trisomy, (2) complex IT involving more than one abnormality, (3) deletion inherited from a parent with a balanced IT resulting in pure segmental monosomy. Of the cases in which follow-up parental studies were available, over half showed inheritance from an apparently unaffected parent carrying the same unbalanced rearrangement detected in the propositi, thus decreasing the likelihood that these IT events are clinically relevant. Nevertheless, we identified six cases in which small submicroscopic events were detected involving known disease-associated genes/genomic segments and are likely to be pathogenic. We recommend that copy number gains detected by clinical aCGH analysis should be confirmed using FISH analysis whenever possible in order to determine the physical location of the duplicated segment. We hypothesize that the increased use of aCGH in the clinic will demonstrate that IT occurs more frequently than previously considered but can identify genomic rearrangements with unclear clinical significance.

FIG. 1

Summary karyogram. An ideogram of each chromosome at an 850-band level resolution (http://www.peds.ufl.edu/divisions/genetics/teaching/chromosomes.htm) is shown with the insertion site on the left indicated with a triangle and the CNV fragment on the right indicated by the horizontal line. The size of the line represents the relative size of the CNV (not drawn to scale). Red represents copy number loss (deletion), green represents copy number gain, and black represents no copy number change (“copy neutral”). Parent of origin is provided when available (U, unbalanced in the parent; B, balanced in the parent; dn, de novo).

FIG. 2

Simple intrachromosomal and interchromosomal insertional translocations. A: Case 1: The aCGH summary table is on top. To the left is the aCGH result showing the oligonucleotide probes specific for chromosome 7 next to the chromosome 7 ideogram. The probes detecting the gain in copy number are shown in green and circled. To the right is the FISH result showing insertion of chromosome 7p14 material (probe RP11-100A8 in red) into 7q. The control probe is RP11-61L9 labeled in green. A partial karyotype of chromosome 7 (G-banded) is shown below. The arrow indicates the derivative chromosome 7 with the box showing the location of the insertion. B: Case 2: The aCGH summary table is on top. On the bottom is the FISH result showing insertion of chromosome Xq material (probe RP11-1145H7 in red) into Xp. The control probe is RP11-383C12 labeled in green. The inset panel is the same X chromosome FISH image shown in inverse DAPI. C: Case 4: The aCGH summary table is on top. To the left is the aCGH result showing the oligonucleotide probes specific for chromosome 1 next to the chromosome 1 ideogram. The probes detecting the gain in copy number are shown in red and circled. To the right is the FISH result showing insertion of chromosome 1q32 material (probe RP11-279E18 in red) into chromosome 5q. The control probe is RP11-339I2 labeled in green. A partial karyotype of chromosome 5 (G-banded) is shown below. The arrow indicates the derivative chromosome 5 with the box showing the location of the insertion. D: Case 25: The aCGH summary table is on top. On the bottom is the inverse DAPI FISH image showing insertion of chromosome 17q material (probes RP11-497H17 in red and RP11-16C1 in green) into chromosome 22p. E: Case 26: The aCGH summary table is on top. On the bottom is the FISH result showing insertion of chromosome 11p material (probe RP11-65C9 in red) into chromosome 22p. The control probe is RP11-709C9 labeled in green. The inset panel is the same chromosome 22 FISH image shown in inverse DAPI.

FIG. 3

Interchromosomal insertions involving similar regions of chromosome 6p material into chromosome 3p. FISH analysis was performed using probe RP1-20B11 (chromosome 6). A: Case 20: To the left is the genome view of the aCGH results with the copy number gain indicated by the red circle. The aCGH summary table is to the right. On the bottom right is the FISH result. The control probe is RP11-239K6 labeled in green. The inset panel is the FISH image shown in inverse DAPI. B: Case 22: The aCGH summary table is on top. On the bottom is the inverse DAPI FISH image for the propositus (left) and the mother (right). The control probe is RP11-69L16 labeled in green.

FIG. 4

Schematic representation of chromosome 6p25 genomic structure. The ideogram for chromosome 6 is shown on top. The red box indicates the region of interest. The enlarged view of the first 1 Mb of chromosome 6p25 is shown below. The blue bar indicates the location of the FISH probe. RefSeq genes are indicated with arrows. The orange bars indicate regions of segmental duplication (Seg Dup). The green bars indicate regions of copy number polymorphisms (CNP).

FIG. 5

Complex insertional translocations. A: Case 29: The aCGH summary table is on top. To the left is the aCGH result showing the oligonucleotide probes specific for chromosome 1 next to the chromosome 1 ideogram. The probes detecting the gain in copy number are shown in green and circled. To the right is the FISH result showing insertion of chromosome 1q32 material (probe RP11-379J10 in red) into chromosome 1p36. The control probe is RP5-888M10 labeled in green, which maps to chromosome 1p36.22. The inset panel is the FISH image shown in inverse DAPI. B: Case 33: The aCGH summary table is on top. On the bottom is the inverse DAPI FISH image showing insertion of chromosome 6q material (probe RP11-211L10 in red) into chromosome Xq. C: Case 34: The aCGH summary table is on top. On the bottom is the FISH image showing insertion of chromosome 5p material (probe RP11-43F13 in green) into chromosome Xq. The X chromosome probes are RP11-119A22 labeled in red and the centromere probe in green (Vysis).

FIG. 6

Deletion resulting from a balanced translocation in a parent (Case 40). A: To the left is the genome view of the aCGH results with the copy number gain indicated by the red circle. The top right is a zoomed in view of the individual oligonucleotide probes detecting the deletion. The bottom right is the deleted region detected on the high-density 244K Agilent array. B: A pedigree of the family. The propositus is indicated by the filled in circle. The father and brother carry the balanced insertional translocation (half-filled squares). Below the pedigree is the FISH image showing the deletion in the propositus (left) and the balanced insertion in the brother (right). The FISH probes are RP11-17O20 in red, which maps to chromosome 21p22.12, and the chromosome 21 telomeric probe (Vysis) in green. The inset panel is the inverse DAPI FISH image of the derivative chromosome 11 in the brother.