Alternative titles; symbols
HGNC Approved Gene Symbol: TOP3A
Cytogenetic location: 17p11.2 Genomic coordinates (GRCh38): 17:18,271,428-18,314,994 (from NCBI)
| Location | Phenotype |
Phenotype MIM number |
Inheritance |
Phenotype mapping key |
|---|---|---|---|---|
| 17p11.2 | Microcephaly, growth restriction, and increased sister chromatid exchange 2 | 618097 | Autosomal recessive | 3 |
| Progressive external ophthalmoplegia with mitochondrial DNA deletions, autosomal recessive 5 | 618098 | Autosomal recessive | 3 |
The TOP3A gene encodes topoisomerase III alpha, which binds to BLM (RECQL3; 604610) as part of the BTRR complex that promotes dissolution of double Holliday junctions (dHJs) that arise as a result of homologous-recombination-mediated repair of double-stranded DNA (dsDNA) breaks during DNA synthesis (summary by Martin et al., 2018).
Mammalian DNA topoisomerase (Topo) III belongs to the type IA DNA Topo subfamily, whose members also include bacterial DNA Topo I and Topo III, and yeast Topo III. S. cerevisiae Topo III may have a significant role in the unlinking of parental strands at the final stage of DNA replication and/or in the dissociation of structures in mitotic cells that could lead to recombination (summary by Li and Wang, 1998).
Through cloning and sequencing, Hanai et al. (1996) identified a human cDNA encoding a protein that is homologous to the Escherichia coli DNA topoisomerase I subfamily of enzymes and that shares no significant sequence homology with eukaryotic DNA topoisomerase I (126420). Expressing the cloned human cDNA in yeast lacking endogenous DNA topoisomerase I yielded an activity in cell extracts that specifically reduced the number of supercoils in a highly negatively supercoiled DNA. On the basis of these results, the human gene containing the cDNAs sequence was denoted TOP3 and the protein it encodes was denoted DNA topoisomerase III by Hanai et al. (1996).
Fritz et al. (1997) isolated a nearly full-length cDNA of human TOP3. Northern blot analysis indicated that TOP3 is expressed in multiple somatic tissues.
Wu et al. (2000) determined that BLM (RECQL3; 604610) and TOP3A colocalized in the nucleus of human cells and coimmunoprecipitated from cell extracts. By in vitro binding assays with truncated BLM mutants, the authors identified 2 independent domains that mediate the interaction with TOP3A. One domain resides between residues 143 and 212 in the N-terminal domain of BLM, and the other resides between residues 1266 and 1417 in the C-terminal domain.
Hu et al. (2001) utilized various deletion constructs of green fluorescent protein (GFP)-tagged BLM to demonstrate that the first 133 amino acids of BLM are necessary and sufficient for interaction between Topo III-alpha and BLM. The Topo III-alpha-interaction domain of BLM is not required for the localization of BLM to the PML nuclear bodies; in contrast, Topo III-alpha is recruited to the PML nuclear bodies via its interaction with BLM. Expression of a full-length BLM (amino acids 1-1417) in Bloom syndrome cells corrected their high sister chromatid exchanges to normal levels, whereas expression of a BLM fragment lacking the Topo III-alpha-interaction domain (amino acids 133-1417) resulted in intermediate sister chromatid exchange levels. The authors hypothesized that the BLM-TOP3A complex is involved in the regulation of recombination in somatic cells.
Wu and Hickson (2003) demonstrated that BLM and TOP3A together effect the resolution of a recombination intermediate containing a double Holliday junction. The mechanism, which they termed double-junction dissolution, is distinct from classical Holliday junction resolution and prevents exchange of flanking sequences. Loss of such an activity explains many of the cellular phenotypes of Bloom syndrome. Wu and Hickson (2003) proposed that double Holliday junctions are formed during the homologous recombination-dependent repair of daughter strand gaps that arise during replication, and that the dissolution of these double Holliday junctions by BLM prevents the diagnostically high sister chromatid exchange frequency seen in Bloom syndrome cells. Furthermore, BLM-catalyzed double-junction dissolution may act to suppress tumorigenesis by preventing loss of heterozygosity, a feature associated with BLM deficiency in mice, through the suppression of ectopic recombination and crossing-over between homologous chromosomes.
Wu et al. (2006) showed that RMI1 (610404) promoted dissolution catalyzed by TOP3A in an vitro assay, but not dissolution catalyzed by other type IA topoisomerases. RMI1 physically interacted with TOP3A, and it appeared to recruit TOP3A to double Holliday junctions. Raynard et al. (2006) showed that RMI1 associated independently with both TOP3A and BLM, and that under physiologic conditions, dissolution of double Holliday junctions by BLM and TOP3A was completely dependent on RMI1.
Takaku et al. (2010) found that human EVL (616912) formed circular heat-stable catemers of single-stranded DNA (ssDNA) in the presence of TOP3A. Catenation depended on the annealing activity of EVL. Surface plasmon resonance analysis confirmed interaction of EVL and TOP3A. Takaku et al. (2010) proposed that EVL and TOP3A may be essential for homologous recombination by processing DNA intermediates formed during this process.
Using gradient separation, Nicholls et al. (2018) found that Top3-alpha was localized to the mitochondrial matrix of HeLa cells. Knockdown of Top3-alpha in HeLa cells resulted in a defect in maintenance and topology of mitochondrial DNA (mtDNA), as mtDNA copy number was depleted and high molecular weight mtDNA species were accumulated. Analysis of the high molecular weight mtDNA species by electron microscopy showed that the loss of Top3-alpha induced the formation of mtDNA catenane. The catenated species appeared as assemblies of circular DNA molecules linked together at a single central point, with only a small proportion of monomeric mtDNA remaining in the Top3-alpha knockdown cells. Catenane formation was centered around the origin of heavy strand (OriH) DNA synthesis region on mitochondrial DNA during the termination of mtDNA replication, forming a structure, termed a hemicatenane by the authors, that consists of a branched molecule with single-strand DNA that does not represent a Holliday junction. Mitochondrial localization of protein components of the BLM-Top3-alpha RMI1-RMI2 (BTR) complex by Western blotting of cell fractions demonstrated that mitochondrial Top3-alpha functions independently of the BTR complex despite nuclear Top3-alpha being a subunit of the BTR complex. Confocal microscopic visualization of the effects of Top3-alpha depletion upon mtDNA separation and segregation showed that Top3-alpha-mediated DNA separation is required for separation of the segregating unit of mtDNA, the nucleoid.
By screening a panel of human/rodent somatic hybrids by Southern analysis and by fluorescence in situ hybridization, Hanai et al. (1996) demonstrated that the human TOP3 gene is present in single copy and is located on 17p12-p11.2.
Microcephaly, Growth Restriction, and Increased Sister Chromatid Exchange 2
In 10 patients from 7 unrelated families with microcephaly, growth restriction, and increased sister chromatid exchange-2 (MGRISCE2; 618097), Martin et al. (2018) identified biallelic mutations in the TOP3A gene (see, e.g., 601243.0001-601243.0003). The mutations, which were found by whole-exome sequencing and confirmed by Sanger sequencing, segregated with the disorder in the families. All except 1 were predicted to result in a frameshift and premature termination, often associated with nonsense-mediated mRNA decay and consistent with a loss of function. One patient (patient 5) was compound heterozygous for a missense mutation (A176V) and a frameshift mutation. Cells derived from several patients showed decreased amounts of TOP3A protein. Patient cells showed significantly increased (3- to 6-fold) more sister chromatid exchanges (SCEs) compared to controls, suggesting extensive crossover recombination. Cells from 1 patient showed chromosome segregation defects during mitosis, with increased chromatin bridges and lagging chromatin and chromosomes, as well as postmitotic defects, such as increased micronuclei and abnormal nuclear bodies. Skeletal muscle biopsy from 1 patient showed significant mitochondrial DNA depletion; similar studies were not available for the other patients. The findings were consistent with an overall accumulation of DNA damage and genomic instability that could impair cell viability during development, causing microcephaly and growth restriction.
Progressive External Ophthalmoplegia With Mitochondrial DNA Deletions, Autosomal Recessive 5
In a 67-year-old woman with autosomal recessive progressive external ophthalmoplegia with mitochondrial DNA deletions-5 (PEOB5; 618098), Nicholls et al. (2018) identified compound heterozygous mutations in the TOP3A gene (M100V, 601243.0004 and R135X, 601243.0005). The mutations, which were found by whole-exome sequencing and confirmed by Sanger sequencing, segregated with the disorder in the family, although DNA from the deceased unaffected father was not available. The R135X variant was predicted to result in a loss of function. In vitro functional expression studies showed that the M100V variant had impaired catalytic activity, and patient cells showed increased levels of catenated mtDNA compared to controls. Patient skeletal muscle biopsy showed numerous COX-deficient ragged-red fibers, increased mtDNA deletions (over 80%), and extensive variable mtDNA rearrangements. The findings illustrated the important role of TOP3A in mtDNA replication and separation.
In 2 sibs with PEOB5, Llaurado et al. (2023) identified a homozygous missense mutation in the TOP3A gene (D205G; 601243.0006). The mutation was identified by sequencing a panel of genes associated with replication and maintenance of mitochondrial DNA and confirmed by Sanger sequencing. Mitochondrial DNA analysis in muscle tissue from one of the sibs demonstrated multiple mtDNA deletions.
TOP3A Deletion in Smith-Magenis Syndrome
Elsea et al. (1998) reported that the TOP3 gene is commonly deleted in patients with the Smith-Magenis syndrome (SMS; 182290) and that the TOP3 gene maps between D17S447 and D17S258 on 17p. The authors concluded that haploinsufficiency of TOP3 does not have a major impact on the behavior of cells from SMS patients and may not play a significant role in the SMS phenotype. In cellular studies of SMS patient lymphoblasts and their respective parental cell lines, Elsea et al. (1998) found that hemizygosity for TOP3 does not affect cell cycle kinetics and/or activation of ionizing radiation-sensitive cell cycle checkpoints. In addition, the induction of apoptosis in response to ionizing radiation in SMS and parental cells was similar.
Li and Wang (1998) carried out targeted disruption of the mouse Top3, or Top3-alpha, gene. No viable homozygous mutant progeny were obtained. Examination of dissected embryos showed that implantation of homozygous mutant embryos and the induction of decidualization could occur, but viability of these embryos was severely compromised at an early stage of development. Heterozygous mutant progeny resembled their wildtype littermates.
In a girl (family 1) with microcephaly, growth restriction, and increased sister chromatid exchange-2 (MGRISCE2; 618097), Martin et al. (2018) identified a homozygous 1-bp deletion (c.2718del, NM_004618.4) in the TOP3A gene, resulting in a frameshift and premature termination (Thr907LeufsTer101). The mutation, which was found by whole-exome sequencing and confirmed by Sanger sequencing, segregated with the disorder in the family. The variant was found at a low frequency (0.00041%) in the gnomAD database. The mutation occurred at the 3-prime end of the gene and was predicted to escape nonsense-mediated mRNA decay and result in the production of a protein lacking the C-terminal zinc finger domain. In vitro functional expression studies showed that the mutation resulted in decreased protein stability compared to wildtype, consistent with loss of enzyme activity being the pathogenic mechanism.
In 6 patients from 4 unrelated families (families 2, 4, 6, and 7) of Middle Eastern descent with microcephaly, growth restriction, and increased sister chromatid exchange-2 (MGRISCE2; 618097), Martin et al. (2018) identified a homozygous 1-bp duplication (c.2271dup, NM_004618.4) in the TOP3A gene, resulting in a frameshift and premature termination (Arg758GlnfsTer3). The mutation, which was found by whole-exome sequencing and confirmed by Sanger sequencing, segregated with the disorder in the families and was not present in the gnomAD database.
In Spanish sibs (family 5) of Middle Eastern descent with microcephaly, growth restriction, and increased sister chromatid exchange-2 (MGRISCE2; 618097), Martin et al. (2018) identified a homozygous 1-bp deletion (c.2428del, NM_004618.4) in the TOP3A gene, resulting in a frameshift and premature termination (Ser810LeufsTer2). The mutation, which was found by whole-exome sequencing and confirmed by Sanger sequencing, segregated with the disorder in the family and was not present in the gnomAD database.
In a 67-year-old woman with autosomal recessive progressive external ophthalmoplegia with mitochondrial DNA deletions-5 (PEOB5; 618098), Nicholls et al. (2018) identified compound heterozygous mutations in the TOP3A gene: a c.298A-G transition (c.298A-G, NM_004618.3), resulting in a met100-to-val (M100V) substitution, and a c.403C-T transition, resulting in an arg135-to-ter (R135X; 601243.0005) substitution. Both mutations occurred in the topoisomerase/primase domain. The mutations, which were found by whole-exome sequencing and confirmed by Sanger sequencing, segregated with the disorder in the family, although DNA from the deceased unaffected father was not available. Both variants were found at low frequencies in heterozygous state in the ExAC database (5.804 x 10(-5) for M100V and 1.983 x 10(-4) for R135X). The R135X variant was predicted to result in a loss of function. In vitro functional expression studies showed that the M100V variant had impaired catalytic activity, and patient cells showed increased levels of catenated mtDNA compared to controls. Patient skeletal muscle biopsy showed numerous COX-deficient ragged-red fibers, increased mtDNA deletions (over 80%), and extensive variable mtDNA rearrangements.
For discussion of the c.403C-T transition (c.403C-T, NM_004618.3) in the TOP3A gene, resulting in an arg135-to-ter (R135X) substitution, that was found in compound heterozygous state in a patient with autosomal recessive progressive external ophthalmoplegia with mitochondrial DNA deletions-5 (PEOB5; 618098) by Nicholls et al. (2018), see 601243.0004.
In 2 sibs with autosomal recessive progressive external ophthalmoplegia with mitochondrial DNA deletions-5 (PEOB5; 618098), Llaurado et al. (2023) identified homozygosity for a c.614A-G transition (c.614A-G, NM_004618.5) in the TOP3A gene, resulting in an asp205-to-gly (D205G) substitution. The mutation, which was identified by sequencing a panel of genes associated with replication and maintenance of mitochondrial DNA and confirmed by Sanger sequencing, segregated with disease in the family. The mutation was not present in the gnomAD database.
Elsea, S. H., Fritz, E., Schoener-Scott, R., Meyn, M. S., Patel, P. I. Gene for topoisomerase III maps within the Smith-Magenis syndrome critical region: analysis of cell-cycle distribution and radiation sensitivity. Am. J. Med. Genet. 75: 104-108, 1998. [PubMed: 9450867]
Fritz, E., Elsea, S. H., Patel, P. I., Meyn, M. S. Overexpression of a truncated human topoisomerase III partially corrects multiple aspects of the ataxia-telangiectasia phenotype. Proc. Nat. Acad. Sci. 94: 4538-4542, 1997. [PubMed: 9114025] [Full Text: https://doi.org/10.1073/pnas.94.9.4538]
Hanai, R., Caron, P. R., Wang, J. C. Human TOP3: a single-copy gene encoding DNA topoisomerase III. Proc. Nat. Acad. Sci. 93: 3653-3657, 1996. [PubMed: 8622991] [Full Text: https://doi.org/10.1073/pnas.93.8.3653]
Hu, P., Beresten, S. F., van Brabant, A. J., Ye, T.-Z., Pandolfi, P.-P., Johnson, F. B., Guarente, L., Ellis, N. A. Evidence for BLM and topoisomerase III-alpha interaction in genomic stability. Hum. Molec. Genet. 10: 1287-1298, 2001. [PubMed: 11406610] [Full Text: https://doi.org/10.1093/hmg/10.12.1287]
Li, W., Wang, J. C. Mammalian DNA topoisomerase III-alpha is essential in early embryogenesis. Proc. Nat. Acad. Sci. 95: 1010-1013, 1998. [PubMed: 9448276] [Full Text: https://doi.org/10.1073/pnas.95.3.1010]
Llaurado, A., Rovira-Moreno, E., Codina-Sola, M., Martinez-Saez, E., Salvado, M., Sanchez-Tejerina, D., Sotoca, J., Lopez-Diego, V., Restrepo-Vera, J. L., Garcia-Arumi, E., Juntas-Morales, R. Chronic progressive external ophthalmoplegia plus syndrome due to homozygous missense variant in TOP3A gene. Clin. Genet. 103: 492-494, 2023. [PubMed: 36544354] [Full Text: https://doi.org/10.1111/cge.14287]
Martin, C.-A., Sarlos, K., Logan, C. V., Thakur, R. S., Parry, D. A., Bizard, A. H., Leitch, A., Cleal, L., Ali, N. S., Al-Owain, M. A., Allen, W., Altmuller, J., and 40 others. Mutations in TOP3A cause a Bloom syndrome-like disorder. Am. J. Hum. Genet. 103: 221-231, 2018. Note: Erratum: Am. J. Hum. Genet. 103: 456 only, 2018. [PubMed: 30057030] [Full Text: https://doi.org/10.1016/j.ajhg.2018.07.001]
Nicholls, T. J., Nadalutti, C. A., Motori, E., Sommerville, E. W., Gorman, G. S., Basu, S., Hoberg, E., Turnbull, D. M., Chinnery, P. F., Larsson, N.-G., Larsson, E., Falkenberg, M., Taylor, R. W., Griffith, J. D., Gustafsson, C. M. Topoisomerase 3-alpha is required for decatenation and segregation of human mtDNA. Molec. Cell 69: 9-23, 2018. [PubMed: 29290614] [Full Text: https://doi.org/10.1016/j.molcel.2017.11.033]
Raynard, S., Bussen, W., Sung, P. A double Holliday junction dissolvasome comprising BLM, topoisomerase III-alpha, and BLAP75. J. Biol. Chem. 281: 13861-13864, 2006. [PubMed: 16595695] [Full Text: https://doi.org/10.1074/jbc.C600051200]
Takaku, M., Takahashi, D., Machida, S., Ueno, H., Hosoya, N., Ikawa, S., Miyagawa, K., Shibata, T., Kurumizaka, H. Single-stranded DNA catenation mediated by human EVL and a type I topoisomerase. Nucleic Acids Res. 38: 7579-7586, 2010. [PubMed: 20639531] [Full Text: https://doi.org/10.1093/nar/gkq630]
Wu, L., Bachrati, C. Z., Ou, J., Xu, C., Yin, J., Chang, M., Wang, W., Li, L., Brown, G. W., Hickson, I. D. BLAP75/RMI1 promotes the BLM-dependent dissolution of homologous recombination intermediates. Proc. Nat. Acad. Sci. 103: 4068-4073, 2006. [PubMed: 16537486] [Full Text: https://doi.org/10.1073/pnas.0508295103]
Wu, L., Davies, S. L., North, P. S., Goulaouic, H., Riou, J.-F., Turley, H., Gatter, K. C., Hickson, I. D. The Bloom's syndrome gene product interacts with topoisomerase III. J. Biol. Chem. 275: 9636-9644, 2000. [PubMed: 10734115] [Full Text: https://doi.org/10.1074/jbc.275.13.9636]
Wu, L., Hickson, I. D. The Bloom's syndrome helicase suppresses crossing over during homologous recombination. Nature 426: 870-874, 2003. [PubMed: 14685245] [Full Text: https://doi.org/10.1038/nature02253]