MRN complex

The MRN complex (MRX complex in yeast) is a protein complex consisting of Mre11, Rad50 and Nbs1 (also known as Nibrin^[1] in humans and as Xrs2 in yeast). In eukaryotes, the MRN/X complex plays an important role in the initial processing of double-strand DNA breaks prior to repair by homologous recombination or non-homologous end joining. The MRN complex binds avidly to double-strand breaks both in vitro and in vivo and may serve to tether broken ends prior to repair by non-homologous end joining or to initiate DNA end resection prior to repair by homologous recombination. The MRN complex also participates in activating the checkpoint kinase ATM in response to DNA damage.^[2][3] Production of short single-strand oligonucleotides by Mre11 endonuclease activity has been implicated in ATM activation by the MRN complex.^[4]

Evolutionary ancestry and biologic function

The MRN complex has been mainly studied in eukaryotes. However, recent work shows that two of the three protein components of this complex, Mre11 and Rad50, are also conserved in extant prokaryotic archaea.^[5] This finding suggests that key components of the eukaryotic MRN complex are derived by evolutionary descent from the archaea. In the archaeon Sulfolobus acidocaldarius, the Mre11 protein interacts with the Rad50 protein and appears to have an active role in the repair of DNA damages experimentally introduced by gamma radiation.^[6] Similarly, during meiosis in the eukaryotic protist Tetrahymena Mre11 is required for repair of DNA damages, in this case double-strand breaks,^[7] by a process that likely involves homologous recombination.

Biological function

Repair of double-strand DNA breaks

In eukaryotes, the MRN complex (through cooperation of its subunits) has been identified as a crucial player in many stages of the repair process of double-strand DNA breaks: initial detection of a lesion, halting of the cell cycle to allow for repair, selection of a specific repair pathway (i.e., via homologous recombination or non-homologous end joining) and providing mechanisms for initiating reconstruction of the DNA molecule (primarily via spatial juxtaposition of the ends of broken chromosomes).^[8] Initial detection is thought to be controlled by both Nbs1 ^[9] and MRE11.^[10] Likewise, cell cycle checkpoint regulation is ultimately controlled by phosphorylation activity of the ATM kinase, which is pathway dependent on both Nbs1 ^[11] and MRE11.^[10] MRE11 alone is known to contribute to repair pathway selection,^[12] while MRE11 and Rad50 work together to spatially align DNA molecules: Rad50 tethers two linear DNA molecules together ^[13] while MRE11 fine-tunes the alignment by binding to the ends of the broken chromosomes.^[14]

Telomere maintenance

Telomeres maintain the integrity of the ends of linear chromosomes during replication and protect them from being recognized as double-strand breaks by the DNA repair machinery. MRN participates in telomere maintenance primarily via association with the TERF2 protein of the shelterin complex.^[15] Additional studies have suggested that Nbs1 is a necessary component protein for telomere elongation by telomerase.^[16] Additionally, knockdown of MRN has been shown to significantly reduce the length of the G-overhang at human telomere ends,^[17] which could inhibit the proper formation of the so-called T-loop, destabilizing the telomere as a whole. Telomere lengthening in cancer cells by the alternative lengthening of telomeres (ALT) mechanism has also been shown to be dependent on MRN, especially on the Nbs1 subunit.^[18] Taken together, these studies suggest MRN plays a crucial role in maintenance of both length and integrity of telomeres.

Role in human disease

Mutations in MRE11 have been identified in patients with an ataxia-telangiectasia-like disorder (ATLD).^[19] Mutations in RAD50 have been linked to a Nijmegen Breakage Syndrome-like disorder (NBSLD).^[20] Mutations in the NBN gene, encoding the human Nbs1 subunit of the MRN complex, are causal for Nijmegen Breakage Syndrome.^[21] All three disorders belong to a group of chromosomal instability syndromes that are associated with impaired DNA damage response and increased cellular sensitivity to ionising radiation.^[22]

Role in human cancer

The MRN complex's roles in cancer development are as varied as its biological functions. Double-strand DNA breaks, which it monitors and signals for repair, may themselves be the cause of carcinogenic genetic alteration,^[23] suggesting MRN provides a protective effect during normal cell homeostasis. However, upregulation of MRN complex sub-units has been documented in certain cancer cell lines when compared to non-malignant somatic cells,^[24] suggesting some cancer cells have developed a reliance on MRN overexpression. Since tumor cells have increased mitotic rates compared to non-malignant cells this is not entirely unexpected, as it is plausible that an increased rate of DNA replication necessitates higher nuclear levels of the MRN complex. However, there is mounting evidence that MRN is itself a component of carcinogenesis, metastasis and overall cancer aggression.

Tumorigenesis

In mice models, mutations in the Nbs1 subunit of MRN alone (producing the phenotypic analog of Nijmegen Breakage Syndrome in humans) have failed to produce tumorigenesis. However, double knockout mice with mutated Nbs1 which were also null of the p53 tumor suppressor gene displayed tumor onset significantly earlier than their p53 wildtype controls.^[25] This implies that Nbs1 mutations are themselves sufficient for tumorigenesis; a lack of malignancy in the control seems attributable to the activity of p53, not of the benignity of Nbs1 mutations. Extension studies have confirmed an increase in B and T-cell lymphomas in Nbs1-mutated mice in conjunction with p53 suppression, indicating potential p53 inactivation in lymphomagenesis,^[26] which occurs more often in NBS patients.^[27][28] Knockdown of MRE11 in various human cancer cell lines has also been associated with a 3-fold increase in the level of p16INK4a tumor suppressor protein,^[29] which is capable of inducing cellular senescence and subsequently halting tumor cell proliferation. This is thought primarily to be the result of methylation of the p16INK4 promotor gene by MRE11. These data suggest maintaining the integrity and normal expression levels of MRN provides a protective effect against tumorigenesis.

Metastasis

Suppression of MRE11 expression in genetically engineered human breast (MCF7) and bone (U2OS) cancer cell lines has resulted in decreased migratory capacity of these cells,^[29] indicating MRN may facilitate metastatic spread of cancer. Decreased expression of MMP-2 and MMP-3 matrix metalloproteinases, which are known to facilitate invasion and metastasis,^[30] occurred concomitantly in these MRE11 knockdown cells. Similarly, overexpression of Nbs1 in human head and neck squamous cell carcinoma (HNSCC) samples has been shown to induce epithelial–mesenchymal transition (EMT), which itself plays a critical role in cancer metastasis.^[31] In this same study, Nbs1 levels were significantly higher in secondary tumor samples than in samples from the primary tumor, providing evidence of a positive correlation between metastatic spread of tumor cells and levels of MRN expression. Taken together, these data suggest at least two of the three subunits of MRN play a role in mediating tumor metastasis, likely via an association between overexpressed MRN and both endogenous (EMT transition) and exogenous (ECM structure) cell migratory mechanisms.

Aggression

Cancer cells almost universally possess upregulated telomere maintenance mechanisms ^[32] which allows for their limitless replicative potential. The MRN complex's biological role in telomere maintenance has prompted research linking MRN to cancer cell immortality. In human HNSCC cell lines, disruption of the Nbs1 gene (which downregulates expression of the entire MRN complex), has resulted in reduced telomere length and persistent lethal DNA damage in these cells.^[33] When combined with treatment of PARP (poly (ADP-ribose) polymerase) inhibitor (known as PARPi), these cells showed an even greater reduction in telomere length, arresting tumor cell proliferation both in vitro and in vivo via mouse models grafted with various HNSCC cell lines. While treatment with PARPi alone has been known to induce apoptosis in BRCA mutated cancer cell lines,^[34] this study shows that MRN downregulation can sensitize BRCA-proficient cells (those not possessing BRCA mutations) to treatment with PARPi, offering an alternative way to control tumor aggression.

The MRN complex has also been implicated in several pathways contributing to the insensitivity of cancer stem cells to the DNA damaging effects of chemotherapy and radiation treatment,^[35] which is a source of overall tumor aggression. Specifically, the MRN inhibitor Mirin (inhibiting MRE11) has been shown to disrupt the ability of ATM kinase to control the G2-M DNA damage checkpoint, which is required for repair of double-strand DNA breaks.^[36] The loss of this checkpoint strips cancer stem cells' ability to repair lethal genetic lesions, making them vulnerable to DNA damaging therapeutic agents. Likewise, overexpression of Nbs1 in HNSCC cells has been correlated with increased activation of the PI3K/AKT pathway, which itself has been shown to contribute to tumor aggression by reducing apoptosis.^[37] Overall, cancer cells appear to rely on MRN's signaling and repair capabilities in response to DNA damage in order to achieve resistance to modern chemo- and radiation therapies.

See also

• Homologous recombination
• MRE11A
• Rad50

References

1. "Atlas of Genetics and Cytogenetics in Oncology and Haematology - NBS1". Retrieved 2008-02-12.
2. Lee, JH; Paull, TT (Apr 2, 2004). "Direct activation of the ATM protein kinase by the Mre11/Rad50/Nbs1 complex". Science. 304 (5667): 93–6. Bibcode:2004Sci...304...93L. doi:10.1126/science.1091496. PMID 15064416. S2CID 26732864.
3. Lee, JH; Paull, TT (Apr 22, 2005). "ATM activation by DNA double-strand breaks through the Mre11-Rad50-Nbs1 complex". Science. 308 (5721): 551–4. Bibcode:2005Sci...308..551L. doi:10.1126/science.1108297. PMID 15790808. S2CID 37711373.
4. Jazayeri A, Balestrini A, Garner E, Haber JE, Costanzo V (2008). "Mre11-Rad50-Nbs1-dependent processing of DNA breaks generates oligonucleotides that stimulate ATM activity". The EMBO Journal. 27 (14): 1953–1962. doi:10.1038/emboj.2008.128. PMC 2453060. PMID 18596698.
5. White, MF (2011). "Homologous recombination in the archaea: the means justify the ends". Biochem Soc Trans. 39 (1): 15–9. doi:10.1042/BST0390015. PMID 21265740.
6. Quaiser, A; Constantinesco, F; White, MF; Forterre, P; Elie, C (2008). "The Mre11 protein interacts with both Rad50 and the HerA bipolar helicase and is recruited to DNA following gamma irradiation in the archaeon Sulfolobus acidocaldarius". BMC Mol Biol. 9: 25. doi:10.1186/1471-2199-9-25. PMC 2288612. PMID 18294364.
7. Lukaszewicz, A; Howard-Till, RA; Novatchkova, M; Mochizuki, K; Loidl, J (2010). "MRE11 and COM1/SAE2 are required for double-strand break repair and efficient chromosome pairing during meiosis of the protist Tetrahymena". Chromosoma. 119 (5): 505–18. doi:10.1007/s00412-010-0274-9. PMID 20422424. S2CID 12642689.
8. Lamarche, BJ; Orazio, NI; Weitzman, MD (10 September 2010). "The MRN complex in double-strand break repair and telomere maintenance". FEBS Letters. 584 (17): 3682–95. doi:10.1016/j.febslet.2010.07.029. PMC 2946096. PMID 20655309.
9. Lukas, Claudia; Falck, Jacob; Bartkova, Jirina; Bartek, Jiri; Lukas, Jiri (24 February 2003). "Distinct spatiotemporal dynamics of mammalian checkpoint regulators induced by DNA damage". Nature Cell Biology. 5 (3): 255–260. doi:10.1038/ncb945. PMID 12598907. S2CID 7061813.
10. Lavin, M F (10 December 2007). "ATM and the Mre11 complex combine to recognize and signal DNA double-strand breaks". Oncogene. 26 (56): 7749–7758. doi:10.1038/sj.onc.1210880. PMID 18066087.
11. You, Z; Chahwan, C; Bailis, J; Hunter, T; Russell, P (July 2005). "ATM activation and its recruitment to damaged DNA require binding to the C terminus of Nbs1". Molecular and Cellular Biology. 25 (13): 5363–79. doi:10.1128/MCB.25.13.5363-5379.2005. PMC 1156989. PMID 15964794.
12. Shibata, A; Moiani, D; Arvai, AS; Perry, J; Harding, SM; Genois, MM; Maity, R; van Rossum-Fikkert, S; Kertokalio, A; Romoli, F; Ismail, A; Ismalaj, E; Petricci, E; Neale, MJ; Bristow, RG; Masson, JY; Wyman, C; Jeggo, PA; Tainer, JA (9 January 2014). "DNA double-strand break repair pathway choice is directed by distinct MRE11 nuclease activities". Molecular Cell. 53 (1): 7–18. doi:10.1016/j.molcel.2013.11.003. PMC 3909494. PMID 24316220.
13. de Jager, M; van Noort, J; van Gent, DC; Dekker, C; Kanaar, R; Wyman, C (November 2001). "Human Rad50/Mre11 is a flexible complex that can tether DNA ends". Molecular Cell. 8 (5): 1129–35. doi:10.1016/s1097-2765(01)00381-1. PMID 11741547.
14. Williams, RS; Moncalian, G; Williams, JS; Yamada, Y; Limbo, O; Shin, DS; Groocock, LM; Cahill, D; Hitomi, C; Guenther, G; Moiani, D; Carney, JP; Russell, P; Tainer, JA (3 October 2008). "Mre11 dimers coordinate DNA end bridging and nuclease processing in double-strand-break repair". Cell. 135 (1): 97–109. doi:10.1016/j.cell.2008.08.017. PMC 2681233. PMID 18854158.
15. Zhu, XD; Küster, B; Mann, M; Petrini, JH; de Lange, T (July 2000). "Cell-cycle-regulated association of RAD50/MRE11/NBS1 with TRF2 and human telomeres". Nature Genetics. 25 (3): 347–52. doi:10.1038/77139. PMID 10888888. S2CID 6689794.
16. Ranganathan, V; Heine, WF; Ciccone, DN; Rudolph, KL; Wu, X; Chang, S; Hai, H; Ahearn, IM; Livingston, DM; Resnick, I; Rosen, F; Seemanova, E; Jarolim, P; DePinho, RA; Weaver, DT (26 June 2001). "Rescue of a telomere length defect of Nijmegen breakage syndrome cells requires NBS and telomerase catalytic subunit". Current Biology. 11 (12): 962–6. doi:10.1016/s0960-9822(01)00267-6. PMID 11448772. S2CID 11631820.
17. Chai, W; Sfeir, AJ; Hoshiyama, H; Shay, JW; Wright, WE (February 2006). "The involvement of the Mre11/Rad50/Nbs1 complex in the generation of G-overhangs at human telomeres". EMBO Reports. 7 (2): 225–30. doi:10.1038/sj.embor.7400600. PMC 1369251. PMID 16374507.
18. Zhong, ZH; Jiang, WQ; Cesare, AJ; Neumann, AA; Wadhwa, R; Reddel, RR (5 October 2007). "Disruption of telomere maintenance by depletion of the MRE11/RAD50/NBS1 complex in cells that use alternative lengthening of telomeres". The Journal of Biological Chemistry. 282 (40): 29314–22. doi:10.1074/jbc.M701413200. PMID 17693401.
19. Stewart GS, Maser RS, Stankovic T, Bressan DA, Kaplan MI, Jaspers NG, Raams A, Byrd PJ, Petrini JH, Taylor AM (1999). "The DNA double-strand break repair gene hMRE11 is mutated in individuals with an ataxia-telangiectasia-like disorder". Cell. 99 (6): 577–87. doi:10.1016/s0092-8674(00)81547-0. PMID 10612394. S2CID 14434655.
20. Waltes R, Kalb R, Gatei M, Kijas AW, Stumm M, Sobeck A, Wieland B, Varon R, Lerenthal Y, Lavin MF, Schindler D, Dörk T (2009). "Human RAD50 deficiency in a Nijmegen Breakage Syndrome-like disorder". Am J Hum Genet. 84 (5): 605–16. doi:10.1016/j.ajhg.2009.04.010. PMC 2681000. PMID 19409520.
21. Varon R, Demuth I, Chrzanowska KH (1993). "Nijmegen Breakage Syndrome". GeneReviews. PMID 20301355.
22. Taylor AM, Rothblum-Oviatt C, Ellis NA, Hickson ID, Meyer S, Crawford TO, Smogorzewska A, Pietrucha B, Weemaes C, Stewart GS (2019). "Chromosome instability syndromes". Nat Rev Dis Primers. 5 (1): 64. doi:10.1038/s41572-019-0113-0. PMC 10617425. PMID 31537806. S2CID 202671095.
23. Czornak, Kamila; Chughtai, Sanaullah; Chrzanowska, Krystyna H. (December 2008). "Mystery of DNA repair: the role of the MRN complex and ATM kinase in DNA damage repair". Journal of Applied Genetics. 49 (4): 383–396. doi:10.1007/BF03195638. PMID 19029686. S2CID 7387378.
24. Kavitha, C.V.; Choudhary, Bibha; Raghavan, Sathees C.; Muniyappa, K. (September 2010). "Differential regulation of MRN (Mre11–Rad50–Nbs1) complex subunits and telomerase activity in cancer cells". Biochemical and Biophysical Research Communications. 399 (4): 575–580. doi:10.1016/j.bbrc.2010.07.117. PMID 20682289.
25. Williams, BR; Mirzoeva, OK; Morgan, WF; Lin, J; Dunnick, W; Petrini, JH (16 April 2002). "A murine model of Nijmegen breakage syndrome". Current Biology. 12 (8): 648–53. doi:10.1016/s0960-9822(02)00763-7. PMID 11967151. S2CID 16989759.
26. Difilippantonio, S; Celeste, A; Fernandez-Capetillo, O; Chen, HT; Reina San Martin, B; Van Laethem, F; Yang, YP; Petukhova, GV; Eckhaus, M; Feigenbaum, L; Manova, K; Kruhlak, M; Camerini-Otero, RD; Sharan, S; Nussenzweig, M; Nussenzweig, A (July 2005). "Role of Nbs1 in the activation of the Atm kinase revealed in humanized mouse models". Nature Cell Biology. 7 (7): 675–85. doi:10.1038/ncb1270. PMID 15965469. S2CID 27172515.
27. Gładkowska-Dura, M; Dzierzanowska-Fangrat, K; Dura, WT; van Krieken, JH; Chrzanowska, KH; van Dongen, JJ; Langerak, AW (November 2008). "Unique morphological spectrum of lymphomas in Nijmegen breakage syndrome (NBS) patients with high frequency of consecutive lymphoma formation". The Journal of Pathology. 216 (3): 337–44. doi:10.1002/path.2418. PMID 18788073. S2CID 20635073.
28. Steffen, J; Maneva, G; Popławska, L; Varon, R; Mioduszewska, O; Sperling, K (15 December 2006). "Increased risk of gastrointestinal lymphoma in carriers of the 657del5 NBS1 gene mutation". International Journal of Cancer. 119 (12): 2970–3. doi:10.1002/ijc.22280. PMID 16998789.
29. Gao, R; Singh, R; Kaul, Z; Kaul, SC; Wadhwa, R (June 2015). "Targeting of DNA Damage Signaling Pathway Induced Senescence and Reduced Migration of Cancer cells". The Journals of Gerontology. Series A, Biological Sciences and Medical Sciences. 70 (6): 701–13. doi:10.1093/gerona/glu019. PMID 24747666.
30. Kessenbrock, K; Plaks, V; Werb, Z (2 April 2010). "Matrix metalloproteinases: regulators of the tumor microenvironment". Cell. 141 (1): 52–67. doi:10.1016/j.cell.2010.03.015. PMC 2862057. PMID 20371345.
31. Voulgari, A; Pintzas, A (December 2009). "Epithelial-mesenchymal transition in cancer metastasis: mechanisms, markers and strategies to overcome drug resistance in the clinic". Biochimica et Biophysica Acta (BBA) - Reviews on Cancer. 1796 (2): 75–90. doi:10.1016/j.bbcan.2009.03.002. PMID 19306912.
32. Reddel, RR (2014). "Telomere maintenance mechanisms in cancer: clinical implications". Current Pharmaceutical Design. 20 (41): 6361–74. doi:10.2174/1381612820666140630101047. PMC 4262939. PMID 24975603.
33. Lajud, SA; Nagda, DA; Yamashita, T; Zheng, J; Tanaka, N; Abuzeid, WM; Civantos, A; Bezpalko, O; O'Malley BW, Jr; Li, D (15 December 2014). "Dual disruption of DNA repair and telomere maintenance for the treatment of head and neck cancer". Clinical Cancer Research. 20 (24): 6465–78. doi:10.1158/1078-0432.CCR-14-0176. PMID 25324139.
34. Farmer, H; McCabe, N; Lord, CJ; Tutt, AN; Johnson, DA; Richardson, TB; Santarosa, M; Dillon, KJ; Hickson, I; Knights, C; Martin, NM; Jackson, SP; Smith, GC; Ashworth, A (14 April 2005). "Targeting the DNA repair defect in BRCA mutant cells as a therapeutic strategy". Nature. 434 (7035): 917–21. Bibcode:2005Natur.434..917F. doi:10.1038/nature03445. PMID 15829967. S2CID 4364706.
35. Skvortsov, S; Debbage, P; Lukas, P; Skvortsova, I (April 2015). "Crosstalk between DNA repair and cancer stem cell (CSC) associated intracellular pathways". Seminars in Cancer Biology. 31: 36–42. doi:10.1016/j.semcancer.2014.06.002. PMID 24954010.
36. Kuroda, S; Urata, Y; Fujiwara, T (2012). "Ataxia-telangiectasia mutated and the Mre11-Rad50-NBS1 complex: promising targets for radiosensitization". Acta Medica Okayama. 66 (2): 83–92. PMID 22525466.
37. Chang, F; Lee, JT; Navolanic, PM; Steelman, LS; Shelton, JG; Blalock, WL; Franklin, RA; McCubrey, JA (March 2003). "Involvement of PI3K/Akt pathway in cell cycle progression, apoptosis, and neoplastic transformation: a target for cancer chemotherapy". Leukemia. 17 (3): 590–603. doi:10.1038/sj.leu.2402824. PMID 12646949.