Mechanisms for Complex Chromosomal Insertions
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE89306 | GEO | 2016/10/30
SECONDARY ACCESSION(S): PRJNA351713
REPOSITORIES: GEO
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Project description:Insertions and deletions (indels) are common sources of structural variation, and insertions originating from spontaneous DNA lesions are frequent in cancer. We developed a highly sensitive assay (Indel-Seq) to monitor rearrangements in human cells at the TRIM37 acceptor locus that reports indels stemming from experimentallyinduced and spontaneous genome instability. Templated insertions, which derive from sequences genome-wide, require contact between donor and acceptor loci, homologous recombination, and are stimulated by DNA end-processing. Insertions are facilitated by transcription and involve a DNA/RNA hybrid intermediate. Indel-Seq reveals that insertions are generated via multiple pathways. The broken acceptor site anneals with a resected DNA break or invades the displaced strand of a transcription bubble or R-loop followed by DNA synthesis, displacement and then ligation by nonhomologous end-joining. Our studies identify transcription-coupled insertions as a critical source of spontaneous genome instability that are distinct from cut-and-paste events
2023-07-03 | GSE234089 | GEO
Project description:Insertions occur when a segment of one chromosome is translocated and inserted into a new region of the same chromosome or a non-homologous chromosome. We report 69 cases with unbalanced insertions identified using array CGH and FISH, representing 1.25% of 5,683 cases referred to our laboratory for array CGH and found to have copy-number abnormalities. Although the majority of insertions were non-recurrent, several recurrent unbalanced insertions were detected, including three der(Y)ins(Y;18)(q?11.2;p11.32p11.32)pat inherited from parents carrying an unbalanced insertion. The clinical significance of these recurrent rearrangements is unclear, although the small size, limited gene content, and inheritance pattern of each suggests the phenotypic consequences may be benign. Moreover, cryptic, submicroscopic duplications were observed at or near the insertion sites in two patients, further confounding the clinical interpretation of these insertions. Using FISH, linear amplification and array CGH we identified a 126-kb duplicated region from 19p13.3 inserted into MECP2 at Xq28 in a patient with symptoms of Rett syndrome. Our results demonstrate that although the interpretation of most non-recurrent insertions is unclear without high-resolution insertion site characterization, the potential for an otherwise benign duplication to result in a clinically relevant outcome through the disruption of a gene necessitates the use of FISH to determine whether copy-number gains detected by array CGH represent tandem duplications or unbalanced insertions. Further follow-up testing using techniques such as linear amplification coupled with array CGH or sequencing should be used to determine gene involvement at the insertion site after FISH has identified the presence of an insertion. 88 total samples analyzed. No replicates included. Each sample run against a standard sex-matched control.
2011-02-28 | E-GEOD-27080 | biostudies-arrayexpress
Project description:A custom high-density oligonucleotide array CGH (HD aCGH) was designed to facilitate the accurate mapping of the boundaries of the recurrent breakpoint at 3q13.31in 8 osteosarcoma tumors and cell lines Keywords: Genetic analysis by HD aCGH
2016-01-01 | GSE12788 | GEO
Project description:Insertions occur when a segment of one chromosome is translocated and inserted into a new region of the same chromosome or a non-homologous chromosome. We report 69 cases with unbalanced insertions identified using array CGH and FISH, representing 1.25% of 5,683 cases referred to our laboratory for array CGH and found to have copy-number abnormalities. Although the majority of insertions were non-recurrent, several recurrent unbalanced insertions were detected, including three der(Y)ins(Y;18)(q?11.2;p11.32p11.32)pat inherited from parents carrying an unbalanced insertion. The clinical significance of these recurrent rearrangements is unclear, although the small size, limited gene content, and inheritance pattern of each suggests the phenotypic consequences may be benign. Moreover, cryptic, submicroscopic duplications were observed at or near the insertion sites in two patients, further confounding the clinical interpretation of these insertions. Using FISH, linear amplification and array CGH we identified a 126-kb duplicated region from 19p13.3 inserted into MECP2 at Xq28 in a patient with symptoms of Rett syndrome. Our results demonstrate that although the interpretation of most non-recurrent insertions is unclear without high-resolution insertion site characterization, the potential for an otherwise benign duplication to result in a clinically relevant outcome through the disruption of a gene necessitates the use of FISH to determine whether copy-number gains detected by array CGH represent tandem duplications or unbalanced insertions. Further follow-up testing using techniques such as linear amplification coupled with array CGH or sequencing should be used to determine gene involvement at the insertion site after FISH has identified the presence of an insertion.
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Project description:Unbalanced translocations are a relatively common type of copy number variation and are a major contributor to neurodevelopmental disorders. We analyzed the breakpoints of 57 unique unbalanced translocations to investigate the mechanisms of how they form. 51 are simple unbalanced translocations between two different chromosome ends, and six rearrangements have more than three breakpoints involving two to five chromosomes. Sequencing 37 breakpoint junctions revealed that simple translocations have between zero and four basepairs (bp) of microhomology (n=26), short inserted sequences (n=8), or paralogous repeats (n=3) at the junctions, indicating that translocations do not arise primarily from non-allelic homologous recombination, but instead form most often via non-homologous end joining or microhomology-mediated break-induced replication. Three simple translocations fuse genes that are predicted to produce in-frame transcripts of SIRPG-WWOX, SMOC2-PROX1, and PIEZO2-MTA1, which may lead to gain of function. Three complex translocations have inversions, insertions, and multiple breakpoint junctions between only two chromosomes. Whole- genome sequencing and fluorescence in situ hybridization analysis of two de novo translocations revealed at least 18 and 33 breakpoints involving five different chromosomes. Breakpoint sequencing of one inherited translocation involving four chromosomes uncovered multiple breakpoints with inversions and insertions. All of these breakpoint junctions had zero to four bp of microhomology consistent with germline chromothripsis, and both de novo events occurred on paternal alleles. Breakpoint sequencing of our large collection of chromosome rearrangements offers a comprehensive analysis of the molecular mechanisms behind germline translocation formation.
2015-04-22 | GSE68019 | GEO
Project description:The high level of human genome structural variation among individuals suggests that there must be portions of the genome that have yet to be discovered, annotated and characterized at the sequence level. Using clone resources developed as part of the Human Genome Structural Variation Sequencing Project, we focused on the characterization of 2,363 novel sequence contigs not present in the human reference genome. We determined that these contigs corresponded to 720 distinct loci of which 400 now have an anchored position in the reference genome. We investigated the sequence properties of these loci and determined that 37% of these novel insertions are copy-number polymorphic. We find that they are significantly enriched within the last 5 Mb of chromosomes (a 2.9-fold enrichment, p=1.0e-18, binomial test) and that most arose as a result of deletions in the human lineage after separation from the African great apes. A subset of these sites shows evidence of marked population stratification among Asian, African and European populations, including a 3.9-kb insertion within the first intron of the lactase gene. Complete sequencing of clones from 192 genomic loci, including 156 completely spanned insertions, provides a detailed and contextual view of 1.67 Mb of inserted sequence. Analysis of this sequence identified 477 elements that show evidence of sequence constraint over evolutionary time, as well as matches to 22 RefSeq gene segments. Twenty-six of the insertions contain matches against mRNA-seq data indicating the potential presence of functionally important, unannotated human sequences. Taking advantage of this high-quality sequence, we develop a method to accurately genotype these novel insertions using next-generation whole-genome sequencing datasets.
2010-04-20 | GSE20634 | GEO
Project description:The noncoding regions of tumor cell genomes harbor a considerable fraction of total variation, but the functional contribution of such variants to tumorigenesis is ill-defined. Among these noncoding variants, somatic insertions are among the least well-characterized due to challenges with interpreting short-read DNA sequences. To address this, we have identified enhancer-associated small insertion variants across a spectrum of tumor cells by using ChIP-seq to enrich and sequence enhancer DNA, and a computational approach with multiple alignment procedures to identify small insertions. Among the 82 tumor genomes studied here, small insertions were frequently observed in enhancer DNA sequences located near known oncogenes. Further study of one such insertion, found in primary leukemia tumor genomes, revealed that it nucleates formation of an active enhancer that drives expression of the LMO2 oncogene. The catalogue of enhancer-associated small insertion variants described here provides a foundation for further investigating the functions of this class of variants across a range of human cancers.
2016-07-08 | GSE76783 | GEO
Project description:Formation of templated insertions at DNA double-strand breaks (DSBs) is very common in cancer cells. The mechanisms and enzymes regulating these events are largely unknown. Here, we investigated templated insertions in yeast at DSBs using amplicon sequencing across a repaired locus. We document very short (most ~5-34 bp), templated inverted duplications at DSBs. They are generated through a foldback mechanism that utilizes microhomologies adjacent to the DSB. Enzymatic requirements suggest a hybrid mechanism wherein one end requires Polδ-mediated synthesis while the other end is captured by nonhomologous end joining (NHEJ) or rarely by alternative end joining (Alt-EJ). This process is exacerbated in mutants with low levels or mutated RPA (rtt105Δ; rfa1-t33) or extensive resection mutant (sgs1Δ exo1Δ). Templated insertions from various distant genomic locations also increase in these mutants as well as in rad27 and originate from fragile regions of the genome. Among complex insertions, common events are insertions of two sequences, originating from the same locus and with inverted orientation. We propose that these inversions are also formed by microhomology-mediated template switching. Taken together, we propose that a shortage of RPA typical in cancer cells is one possible factor stimulating the formation of templated insertions.
2024-03-07 | GSE260753 | GEO
Project description:Unbalanced translocations are a relatively common type of copy number variation and are a major contributor to neurodevelopmental disorders. We analyzed the breakpoints of 57 unique unbalanced translocations to investigate the mechanisms of how they form. 51 are simple unbalanced translocations between two different chromosome ends, and six rearrangements have more than three breakpoints involving two to five chromosomes. Sequencing 37 breakpoint junctions revealed that simple translocations have between zero and four basepairs (bp) of microhomology (n=26), short inserted sequences (n=8), or paralogous repeats (n=3) at the junctions, indicating that translocations do not arise primarily from non-allelic homologous recombination, but instead form most often via non-homologous end joining or microhomology-mediated break-induced replication. Three simple translocations fuse genes that are predicted to produce in-frame transcripts of SIRPG-WWOX, SMOC2-PROX1, and PIEZO2-MTA1, which may lead to gain of function. Three complex translocations have inversions, insertions, and multiple breakpoint junctions between only two chromosomes. Whole- genome sequencing and fluorescence in situ hybridization analysis of two de novo translocations revealed at least 18 and 33 breakpoints involving five different chromosomes. Breakpoint sequencing of one inherited translocation involving four chromosomes uncovered multiple breakpoints with inversions and insertions. All of these breakpoint junctions had zero to four bp of microhomology consistent with germline chromothripsis, and both de novo events occurred on paternal alleles. Breakpoint sequencing of our large collection of chromosome rearrangements offers a comprehensive analysis of the molecular mechanisms behind germline translocation formation. High resolution array CGH; two-color experiment, clinical patient vs. normal control gDNA; sex mis-matched
2015-04-22 | E-GEOD-68019 | biostudies-arrayexpress
Project description:The Complex Architecture of Transgene Insertions
| PRJNA427134 | ENA