Microhomology-mediated end joining

Microhomology-mediated end joining (MMEJ), also known as alternative nonhomologous end-joining (Alt-NHEJ) is one of the pathways for repairing double-strand breaks in DNA. As reviewed by McVey and Lee,^[1] the foremost distinguishing property of MMEJ is the use of microhomologous sequences during the alignment of broken ends before joining, thereby resulting in deletions flanking the original break. MMEJ is frequently associated with chromosome abnormalities such as deletions, translocations, inversions and other complex rearrangements.

There are multiple pathways for repairing double strand breaks, mainly non-homologous end joining (NHEJ), homologous recombination (HR), and MMEJ. NHEJ directly joins both ends of the double strand break and is relatively accurate, although small (usually less than a few nucleotides) insertions or deletions sometimes occur. HR is highly accurate and uses the sister chromatid as a template for accurate repair of the DSB. MMEJ is distinguished from these other repair mechanisms by its use of microhomologous sequences to align the broken strands. This results in frequent deletions and occasionally insertions which are much larger than those produced by NHEJ ^{[citation needed]}. MMEJ is completely independent from classical NHEJ and does not rely on NHEJ core factors such as Ku protein, DNA-PK, or Ligase IV.^[2]

In MMEJ, repair of the DSB is initiated by end resection by the MRE nuclease, leaving single stranded overhangs.^[3] These single stranded overhangs anneal at microhomologies, which are short regions of complementarity, often 5–25 base pairs, between the two strands. A specialized form of MMEJ, called polymerase theta-mediated end-joining (TMEJ), is able to repair breaks using ≥1 bp of homology.^[4]^[5] The helicase domain of DNA polymerase theta possesses ATP-dependent single-strand annealing activity and may promote annealing of microhomologies.^[6] Following annealing, any overhanging bases (flaps) are removed by nucleases such as Fen1 and gaps are filled in by DNA polymerase theta.^[7] This gap-filling ability of polymerase theta helps to stabilize the annealing of ends with minimal complementarity. Besides microhomology footprints, polymerase theta's mutational signature also consists of (infrequent) templated inserts, which are thought to be the result of aborted template-dependent extension, followed by re-annealing at secondary homologous sequences.^[5]

Cell cycle regulation

MMEJ repair is low in G0/G1 phase but is increased during S-phase and G2 phase of the cell cycle.^[3] In contrast, NHEJ operates throughout the cell cycle, and homologous recombination (HR) operates only in late S and G2.

Double strand break repair pathway choice

The choice of which pathway is used for double strand break repair is complex. In most cases, MMEJ accounts for a minor proportion (10%) of double strand break repair, most likely in cases where the double strand break is resected but a sister chromatid is not available for homologous recombination.^[3] Cells which are deficient in either classical NHEJ or HR typically display increased MMEJ. Human homologous recombination factors suppress mutagenic MMEJ following double-strand break resection.^[8]

Genes required

A biochemical assay system shows that at least 6 genes are required for microhomology-mediated end joining: FEN1, Ligase III, MRE11, NBS1, PARP1 and XRCC1.^[9] All six of these genes are up-regulated in one or more cancers. In humans, DNA polymerase theta, encoded by the POLQ gene, plays a central role in microhomology-mediated end joining.^[7] Polymerase theta utilizes its helicase domain to displace replication protein A (RPA) from DNA ends and promote microhomology annealing.^[6] Polymerase theta also uses its polymerase activity to conduct fill-in synthesis, which helps to stabilize paired ends.

Helicase Q, which is conserved in humans, is necessary for polymerase theta-independent MMEJ, as shown using mutational footprint analyses in Caenorrhabditis elegans.^[10]

In cancer

Approximately half of all ovarian cancers are deficient in homologous recombination (HR). These HR-deficient tumors upregulate polymerase theta (POLQ), resulting in an increase in MMEJ.^[11] These tumors are hyper-reliant upon MMEJ, so that knockdown of polymerase theta results in substantial lethality. In most cell types, MMEJ makes a minor contribution to double strand break repair. The hyper-reliance of HR-deficient tumors upon MMEJ may represent a possible drug target for cancer treatment.

MMEJ always involves insertions or deletions, so that it is a mutagenic pathway.^[12] Cells with increased MMEJ may have higher genomic instability and a predisposition towards cancer development, although this has not been demonstrated directly.

In a crustacean

Penaeus monodon is a marine crustacean widely consumed for its nutritional value. Repair of double-strand breaks in this organism can occur by HRR, but NHEJ is undetectable.^[13] While HRR appears to be the major double-strand break repair pathway, MMEJ was also found to play a significant role in repair of DNA double-strand breaks.^[13]

References

^ McVey M, Lee SE (November 2008). "MMEJ repair of double-strand breaks (director's cut): deleted sequences and alternative endings". Trends in Genetics. 24 (11): 529–538. doi:10.1016/j.tig.2008.08.007. PMC 5303623. PMID 18809224.
^ Simsek D, Jasin M (April 2010). "Alternative end-joining is suppressed by the canonical NHEJ component Xrcc4-ligase IV during chromosomal translocation formation". Nature Structural & Molecular Biology. 17 (4): 410–416. doi:10.1038/nsmb.1773. PMC 3893185. PMID 20208544.
^ ^a ^b ^c Truong LN, Li Y, Shi LZ, Hwang PY, He J, Wang H, et al. (May 2013). "Microhomology-mediated End Joining and Homologous Recombination share the initial end resection step to repair DNA double-strand breaks in mammalian cells". Proceedings of the National Academy of Sciences of the United States of America. 110 (19): 7720–7725. Bibcode:2013PNAS..110.7720T. doi:10.1073/pnas.1213431110. PMC 3651503. PMID 23610439.
^ Roerink SF, van Schendel R, Tijsterman M (June 2014). "Polymerase theta-mediated end joining of replication-associated DNA breaks in C. elegans". Genome Research. 24 (6): 954–962. doi:10.1101/gr.170431.113. PMC 4032859. PMID 24614976.
^ ^a ^b Schimmel J, van Schendel R, den Dunnen JT, Tijsterman M (September 2019). "Templated Insertions: A Smoking Gun for Polymerase Theta-Mediated End Joining". Trends in Genetics. 35 (9): 632–644. doi:10.1016/j.tig.2019.06.001. PMID 31296341. S2CID 195892718.
^ ^a ^b Mateos-Gomez PA, Kent T, Deng SK, McDevitt S, Kashkina E, Hoang TM, et al. (December 2017). "The helicase domain of Polθ counteracts RPA to promote alt-NHEJ". Nature Structural & Molecular Biology. 24 (12): 1116–1123. doi:10.1038/nsmb.3494. PMC 6047744. PMID 29058711.
^ ^a ^b Sfeir A, Symington LS (November 2015). "Microhomology-Mediated End Joining: A Back-up Survival Mechanism or Dedicated Pathway?". Trends in Biochemical Sciences. 40 (11): 701–714. doi:10.1016/j.tibs.2015.08.006. PMC 4638128. PMID 26439531.
^ Ahrabi S, Sarkar S, Pfister SX, Pirovano G, Higgins GS, Porter AC, Humphrey TC (July 2016). "A role for human homologous recombination factors in suppressing microhomology-mediated end joining". Nucleic Acids Research. 44 (12): 5743–5757. doi:10.1093/nar/gkw326. PMC 4937322. PMID 27131361.
^ Sharma S, Javadekar SM, Pandey M, Srivastava M, Kumari R, Raghavan SC (March 2015). "Homology and enzymatic requirements of microhomology-dependent alternative end joining". Cell Death & Disease. 6 (3): e1697. doi:10.1038/cddis.2015.58. PMC 4385936. PMID 25789972.
^ Kamp JA, Lemmens BB, Romeijn RJ, Changoer SC, van Schendel R, Tijsterman M (December 2021). "Helicase Q promotes homology-driven DNA double-strand break repair and prevents tandem duplications". Nature Communications. 12 (1): 7126. Bibcode:2021NatCo..12.7126K. doi:10.1038/s41467-021-27408-z. PMC 8654963. PMID 34880204.
^ Ceccaldi R, Liu JC, Amunugama R, Hajdu I, Primack B, Petalcorin MI, et al. (February 2015). "Homologous-recombination-deficient tumours are dependent on Polθ-mediated repair". Nature. 518 (7538): 258–262. Bibcode:2015Natur.518..258C. doi:10.1038/nature14184. PMC 4415602. PMID 25642963.
^ Liang L, Deng L, Chen Y, Li GC, Shao C, Tischfield JA (September 2005). "Modulation of DNA end joining by nuclear proteins". The Journal of Biological Chemistry. 280 (36): 31442–31449. doi:10.1074/jbc.M503776200. PMID 16012167.
^ ^a ^b Srivastava S, Dahal S, Naidu SJ, Anand D, Gopalakrishnan V, Kooloth Valappil R, Raghavan SC (April 2017). "DNA double-strand break repair in Penaeus monodon is predominantly dependent on homologous recombination". DNA Research. 24 (2): 117–128. doi:10.1093/dnares/dsw059. PMC 5397610. PMID 28431013.

General references

[pmid18809224-1] McVey M, Lee SE (November 2008). "MMEJ repair of double-strand breaks (director's cut): deleted sequences and alternative endings". Trends in Genetics. 24 (11): 529–538. doi:10.1016/j.tig.2008.08.007. PMC 5303623. PMID 18809224.

[2] Simsek D, Jasin M (April 2010). "Alternative end-joining is suppressed by the canonical NHEJ component Xrcc4-ligase IV during chromosomal translocation formation". Nature Structural & Molecular Biology. 17 (4): 410–416. doi:10.1038/nsmb.1773. PMC 3893185. PMID 20208544.

[pmid23610439-3] Truong LN, Li Y, Shi LZ, Hwang PY, He J, Wang H, et al. (May 2013). "Microhomology-mediated End Joining and Homologous Recombination share the initial end resection step to repair DNA double-strand breaks in mammalian cells". Proceedings of the National Academy of Sciences of the United States of America. 110 (19): 7720–7725. Bibcode:2013PNAS..110.7720T. doi:10.1073/pnas.1213431110. PMC 3651503. PMID 23610439.

[4] Roerink SF, van Schendel R, Tijsterman M (June 2014). "Polymerase theta-mediated end joining of replication-associated DNA breaks in C. elegans". Genome Research. 24 (6): 954–962. doi:10.1101/gr.170431.113. PMC 4032859. PMID 24614976.

[Schimmel_632–644-5] Schimmel J, van Schendel R, den Dunnen JT, Tijsterman M (September 2019). "Templated Insertions: A Smoking Gun for Polymerase Theta-Mediated End Joining". Trends in Genetics. 35 (9): 632–644. doi:10.1016/j.tig.2019.06.001. PMID 31296341. S2CID 195892718.

[The_helicase_domain_of_Polθ_counter-6] Mateos-Gomez PA, Kent T, Deng SK, McDevitt S, Kashkina E, Hoang TM, et al. (December 2017). "The helicase domain of Polθ counteracts RPA to promote alt-NHEJ". Nature Structural & Molecular Biology. 24 (12): 1116–1123. doi:10.1038/nsmb.3494. PMC 6047744. PMID 29058711.

[Microhomology-Mediated_End_Joining-7] Sfeir A, Symington LS (November 2015). "Microhomology-Mediated End Joining: A Back-up Survival Mechanism or Dedicated Pathway?". Trends in Biochemical Sciences. 40 (11): 701–714. doi:10.1016/j.tibs.2015.08.006. PMC 4638128. PMID 26439531.

[pmid27131361-8] Ahrabi S, Sarkar S, Pfister SX, Pirovano G, Higgins GS, Porter AC, Humphrey TC (July 2016). "A role for human homologous recombination factors in suppressing microhomology-mediated end joining". Nucleic Acids Research. 44 (12): 5743–5757. doi:10.1093/nar/gkw326. PMC 4937322. PMID 27131361.

[pmid25789972-9] Sharma S, Javadekar SM, Pandey M, Srivastava M, Kumari R, Raghavan SC (March 2015). "Homology and enzymatic requirements of microhomology-dependent alternative end joining". Cell Death & Disease. 6 (3): e1697. doi:10.1038/cddis.2015.58. PMC 4385936. PMID 25789972.

[10] Kamp JA, Lemmens BB, Romeijn RJ, Changoer SC, van Schendel R, Tijsterman M (December 2021). "Helicase Q promotes homology-driven DNA double-strand break repair and prevents tandem duplications". Nature Communications. 12 (1): 7126. Bibcode:2021NatCo..12.7126K. doi:10.1038/s41467-021-27408-z. PMC 8654963. PMID 34880204.

[11] Ceccaldi R, Liu JC, Amunugama R, Hajdu I, Primack B, Petalcorin MI, et al. (February 2015). "Homologous-recombination-deficient tumours are dependent on Polθ-mediated repair". Nature. 518 (7538): 258–262. Bibcode:2015Natur.518..258C. doi:10.1038/nature14184. PMC 4415602. PMID 25642963.

[Liang-12] Liang L, Deng L, Chen Y, Li GC, Shao C, Tischfield JA (September 2005). "Modulation of DNA end joining by nuclear proteins". The Journal of Biological Chemistry. 280 (36): 31442–31449. doi:10.1074/jbc.M503776200. PMID 16012167.

[pmid28431013-13] Srivastava S, Dahal S, Naidu SJ, Anand D, Gopalakrishnan V, Kooloth Valappil R, Raghavan SC (April 2017). "DNA double-strand break repair in Penaeus monodon is predominantly dependent on homologous recombination". DNA Research. 24 (2): 117–128. doi:10.1093/dnares/dsw059. PMC 5397610. PMID 28431013.

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Microhomology-mediated end joining

Cell cycle regulation

Double strand break repair pathway choice

Genes required

In cancer

In a crustacean

References

General references

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