Comprehensive analysis of F8 large deletions: Characterization of full breakpoint junctions and description of a possible DNA breakage hotspot in intron 6.
F8
Alu elements
factor VIII
hemophilia A
sequence deletion
Journal
Journal of thrombosis and haemostasis : JTH
ISSN: 1538-7836
Titre abrégé: J Thromb Haemost
Pays: England
ID NLM: 101170508
Informations de publication
Date de publication:
10 2022
10 2022
Historique:
revised:
20
07
2022
received:
18
03
2022
accepted:
25
07
2022
pubmed:
28
7
2022
medline:
23
9
2022
entrez:
27
7
2022
Statut:
ppublish
Résumé
Large F8 deletions represent 3-5% of the variations found in severe hemophilia A patients, but only a few deletion breakpoints have been characterized precisely. Resolving at the nucleotide level 24 F8 large deletions to provide new data on the mechanisms involved in these rearrangements. Breakpoint junctions of 24 F8 large deletions were characterized using a combination of long-range polymerase chain reaction, whole F8 NGS sequencing, and Sanger sequencing. Repeat elements, non-B DNA, and secondary structures were analyzed around the breakpoints. Deletions ranged from 1.667 kb to 0.5 Mb in size. Nine involved F8 neighboring genes. Simple blunt ends and 2-4 bp microhomologies were identified at the breakpoint junctions of 10 (42%) and 8 (33%) deletions, respectively. Five (21%) deletions resulted from homeologous recombination between two Alu elements. The remaining case corresponded to a more complex rearrangement with an insertion of a 19 bp-inverted sequence at the junction. Four different breakpoints were located in a 562-bp region in F8 intron 6. This finding suggested that this region, composed of two Alu elements, is a DNA breakage hotspot. Non-B DNA and secondary structures were identified in the junction regions and may contribute to DNA breakage. Molecular characterization of deletion breakpoints revealed that non-homologous non-replicative DNA repair mechanisms and replication-based mechanisms seemed to be the main causative mechanisms of F8 large deletions. Moreover, we identified a possible F8 DNA breakage hotspot involved in non-recurrent rearrangements.
Sections du résumé
BACKGROUND
Large F8 deletions represent 3-5% of the variations found in severe hemophilia A patients, but only a few deletion breakpoints have been characterized precisely.
OBJECTIVES
Resolving at the nucleotide level 24 F8 large deletions to provide new data on the mechanisms involved in these rearrangements.
METHODS
Breakpoint junctions of 24 F8 large deletions were characterized using a combination of long-range polymerase chain reaction, whole F8 NGS sequencing, and Sanger sequencing. Repeat elements, non-B DNA, and secondary structures were analyzed around the breakpoints.
RESULTS
Deletions ranged from 1.667 kb to 0.5 Mb in size. Nine involved F8 neighboring genes. Simple blunt ends and 2-4 bp microhomologies were identified at the breakpoint junctions of 10 (42%) and 8 (33%) deletions, respectively. Five (21%) deletions resulted from homeologous recombination between two Alu elements. The remaining case corresponded to a more complex rearrangement with an insertion of a 19 bp-inverted sequence at the junction. Four different breakpoints were located in a 562-bp region in F8 intron 6. This finding suggested that this region, composed of two Alu elements, is a DNA breakage hotspot. Non-B DNA and secondary structures were identified in the junction regions and may contribute to DNA breakage.
CONCLUSION
Molecular characterization of deletion breakpoints revealed that non-homologous non-replicative DNA repair mechanisms and replication-based mechanisms seemed to be the main causative mechanisms of F8 large deletions. Moreover, we identified a possible F8 DNA breakage hotspot involved in non-recurrent rearrangements.
Identifiants
pubmed: 35894111
doi: 10.1111/jth.15835
pii: S1538-7836(22)19061-6
doi:
Substances chimiques
Nucleotides
0
DNA
9007-49-2
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
2293-2305Informations de copyright
© 2022 International Society on Thrombosis and Haemostasis.
Références
White GC II, Rosendaal F, Aledort LM, et al. Definitions in hemophilia. Recommendation of the scientific subcommittee on factor VIII and factor IX of the scientific and standardization committee of the International Society on Thrombosis and Haemostasis. Thromb Haemost. 2001;85:560.
Carvalho CMB, Lupski JR. Mechanisms underlying structural variant formation in genomic disorders. Nat Rev Genet. 2016;17:224-238.
Lee JA, Carvalho CMB, Lupski JR. A DNA replication mechanism for generating nonrecurrent rearrangements associated with genomic disorders. Cell. 2007;131:1235-1247.
Bagnall RD, Giannelli F, Green PM. Int22h-related inversions causing hemophilia A: a novel insight into their origin and a new more discriminant PCR test for their detection. J Thromb Haemost. 2006;4:591-598.
Bagnall RD, Giannelli F, Green PM. Polymorphism and hemophilia A causing inversions in distal Xq28: a complex picture. J Thromb Haemost. 2005;3:2598-2599.
Antonarakis SE, Rossiter JP, Young M, et al. Factor VIII gene inversions in severe hemophilia A: results of an international consortium study. Blood. 1995;86:2206-2212.
Bagnall RD, Waseem N, Green PM, Giannelli F. Recurrent inversion breaking intron 1 of the factor VIII gene is a frequent cause of severe hemophilia A. Blood. 2002;99:168-174.
Lannoy N, Hermans C. Principles of genetic variations and molecular diseases: applications in hemophilia A. Crit Rev Oncol Hematol. 2016;104:1-8.
Abelleyro MM, Radic CP, Marchione VD, et al. Molecular insights into the mechanism of nonrecurrent F8 structural variants: Full breakpoint characterization and bioinformatics of DNA elements implicated in the upmost severe phenotype in hemophilia A. Hum Mutat. 2020;41:825-836.
You GL, Ding QL, Lu YL, et al. Characterization of large deletions in the F8 gene using multiple competitive amplification and the genome walking technique. J Thromb Haemost. 2013;11:1103-1110.
Roth L, Marschalek R, Oldenburg J, Oyen F, Schneppenheim R. Characterisation of two novel large F8 deletions in patients with severe haemophilia A and factor VIII inhibitors. Thromb Haemost. 2011;105:279-284.
Vidal F, Farssac E, Tusell J, Puig L, Gallardo D. First molecular characterization of an unequal homologous alu-mediated recombination event responsible for hemophilia. Thromb Haemost. 2002;88:12-16.
Rossetti LC, Goodeve A, Larripa IB, De Brasi CD. Homeologous recombination between AluSx-sequences as a cause of hemophilia. Hum Mutat. 2004;24:440.
Jourdy Y, Chatron N, Fretigny M, et al. Molecular cytogenetic characterization of five F8 complex rearrangements: utility for haemophilia A genetic counselling. Haemophilia. 2017;23:e316-e323.
Li H. Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM. ArXiv:1303.3997 Q-Bio 2013.
Kent WJ. BLAT--the BLAST-like alignment tool. Genome Res. 2002;12:656-664.
Jourdy Y, Bardel C, Fretigny M, et al. Complete characterisation of two new large Xq28 duplications involving F8 using whole genome sequencing in patients without haemophilia A. Haemophilia. 2021;1:117-124.
Oldenburg J, Pavlova A. Genetic risk factors for inhibitors to factors VIII and IX. Haemophilia. 2006;12(Suppl 6):15-22.
Miskinyte S, Butler MG, Hervé D, et al. Loss of BRCC3 deubiquitinating enzyme leads to abnormal angiogenesis and is associated with syndromic moyamoya. Am J Hum Genet. 2011;88:718-728.
Janczar S, Fogtman A, Koblowska M, et al. Novel severe hemophilia A and moyamoya (SHAM) syndrome caused by Xq28 deletions encompassing F8 and BRCC3 genes. Blood. 2014;123:4002-4004.
Fujita J, Miyawaki Y, Suzuki A, et al. A possible mechanism for Inv22-related F8 large deletions in severe hemophilia A patients with high responding factor VIII inhibitors. J Thromb Haemost. 2012;10:2099-2107.
El-Hattab AW, Fang P, Jin W, et al. Int22h-1/int22h-2-mediated Xq28 rearrangements: intellectual disability associated with duplications and in utero male lethality with deletions. J Med Genet. 2011;48:840-850.
Lander ES, Linton LM, Birren B, et al. Initial sequencing and analysis of the human genome. Nature. 2001;409:860-921.
Cordaux R, Batzer MA. The impact of retrotransposons on human genome evolution. Nat Rev Genet. 2009;10:691-703.
Payer LM, Burns KH. Transposable elements in human genetic disease. Nat Rev Genet. 2019;20:760-772.
Ade C, Roy-Engel AM, Deininger PL. Alu elements: an intrinsic source of human genome instability. Curr Opin Virol. 2013;3:639-645.
Boone PM, Yuan B, Campbell IM, et al. The Alu-Rich genomic architecture of SPAST predisposes to diverse and functionally distinct disease-associated CNV alleles. Am J Hum Genet. 2014;95:143-161.
Gu S, Yuan B, Campbell IM, et al. Alu-mediated diverse and complex pathogenic copy-number variants within human chromosome 17 at p13.3. Hum Mol Genet. 2015;24:4061-4077.
White TB, Morales ME, Deininger PL. Alu elements and DNA double-strand break repair. Mob Genet Elem. 2015;5:81-85.
Flynn EK, Kamat A, Lach FP, et al. Comprehensive analysis of pathogenic deletion variants in Fanconi anemia genes. Hum Mutat. 2014;35:1342-1353.
Zimmermann MA, Oldenburg J, Müller CR, Rost S. Characterization of duplication breakpoints in the factor VIII gene. J Thromb Haemost. 2010;8:2696-2704.
Stoppa-Lyonnet D, Duponchel C, Meo T, et al. Recombinational biases in the rearranged C1-inhibitor genes of hereditary angioedema patients. Am J Hum Genet. 1991;49:1055-1062.
Vocke CD, Ricketts CJ, Schmidt LS, et al. Comprehensive characterization of Alu-mediated breakpoints in germline VHL gene deletions and rearrangements in patients from 71 VHL families. Hum Mutat. 2021;42:520-529.
Wang G, Vasquez KM. Impact of alternative DNA structures on DNA damage, DNA repair, and genetic instability. DNA Repair. 2014;19:143-151.
Geng C, Tong Y, Zhang S, et al. Sequence and structure characteristics of 22 deletion breakpoints in intron 44 of the DMD gene based on long-read sequencing. Front Genet. 2021;12:638220.
Summerer A, Mautner V-F, Upadhyaya M, et al. Extreme clustering of type-1 NF1 deletion breakpoints co-locating with G-quadruplex forming sequences. Hum Genet. 2018;137:511-520.
Wu X, Lu Y, Ding Q, et al. Characterisation of large F9 deletions in seven unrelated patients with severe haemophilia B. Thromb Haemost. 2014;112:459-465.