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Histochemistry and Cell Biology

, Volume 141, Issue 1, pp 101–109 | Cite as

RNA-binding protein RBM8A (Y14) and MAGOH localize to centrosome in human A549 cells

  • Yasuhito IshigakiEmail author
  • Yuka Nakamura
  • Takanori Tatsuno
  • Mitsumasa Hashimoto
  • Kuniyoshi Iwabuchi
  • Naohisa TomosugiEmail author
Original Paper

Abstract

RBM8A (Y14) is carrying RNA-binding motif and forms the tight heterodimer with MAGOH. The heterodimer is known to be a member of exon junction complex on exporting mRNA and is required for mRNA metabolisms such as splicing, mRNA export and nonsense-mediated mRNA decay. Almost all RBM8A–MAGOH complexes localize in nucleoplasm and shuttle between nuclei and cytoplasm for RNA metabolism. Recently, the abnormality of G2/M transition and aberrant centrosome regulation in RBM8A- or MAGOH-deficient cells has been reported. These results prompt us to the reevaluation of the localization of RBM8A–MAGOH in human cells. Interestingly, our immunostaining experiments showed the localization of these proteins in centrosome in addition to nuclei. Furthermore, the transiently expressed eYFP-tagged RBM8A and Flag-tagged MAGOH also co-localized with centrosome signals. In addition, the proximity ligation in situ assay was performed to detect the complex formation in centrosome. Our experiments clearly showed that Myc-tagged RBM8A and Flag-tagged MAGOH formed a complex in centrosome. GFP-tagged PLK1 also co-localized with Myc-RBM8A. Our results show that RBM8A–MAGOH complex is required for M-phase progression via direct localization to centrosome rather than indirect effect.

Keywords

RBM8A (Y14) MAGOH Cell cycle Centrosome 

Notes

Acknowledgments

Authors also thank to Prof. Tsutomu Takegami, Prof. Hideaki Nakagawa and Ms. Yoshie Yoshida at the Kanazawa Medical University for their kind support. This work was supported by Grants from Kanazawa Medical University (S2012-1, SR2012-02), Sumitomo Science Foundation, Grants of Strategic Research Project (H2012-16[S1201022]) from Kanazawa Medical University and Ministry of Education, Culture, Sports, Science and Technology, Japan and Grant-in-Aid for Scientific Research in Japan (KAKENHI #25460376).

Conflict of interest

The authors have read the journal’s policy and have the following conflicts: Prof. Naohisa Tomosugi is the president of Medical Care Proteomics Biotechnology Co., Ltd. This does not alter the author’s adherence to all the policies on sharing data.

Supplementary material

418_2013_1135_MOESM1_ESM.docx (353 kb)
Supplementary material 1 (DOCX 352 kb)

References

  1. Aizer A, Brody Y, Ler LW, Sonenberg N, Singer RH, Shav-Tal Y (2008) The dynamics of mammalian P body transport, assembly, and disassembly in vivo. Mol Biol Cell 19(10):4154–4166. doi: 10.1091/mbc.E08-05-0513 PubMedCentralPubMedCrossRefGoogle Scholar
  2. Albers CA, Paul DS, Schulze H, Freson K, Stephens JC, Smethurst PA, Jolley JD, Cvejic A, Kostadima M, Bertone P, Breuning MH, Debili N, Deloukas P, Favier R, Fiedler J, Hobbs CM, Huang N, Hurles ME, Kiddle G, Krapels I, Nurden P, Ruivenkamp CA, Sambrook JG, Smith K, Stemple DL, Strauss G, Thys C, van Geet C, Newbury-Ecob R, Ouwehand WH, Ghevaert C (2012) Compound inheritance of a low-frequency regulatory SNP and a rare null mutation in exon-junction complex subunit RBM8A causes TAR syndrome. Nat Genet 44(4):435–439, S431–S432. doi: 10.1038/ng.1083 Google Scholar
  3. Alliegro MC, Alliegro MA, Palazzo RE (2006) Centrosome-associated RNA in surf clam oocytes. Proc Natl Acad Sci USA 103(24):9034–9038. doi: 10.1073/pnas.0602859103 PubMedCrossRefGoogle Scholar
  4. Bettencourt-Dias M, Glover DM (2007) Centrosome biogenesis and function: centrosomics brings new understanding. Nat Rev Mol Cell Biol 8(6):451–463. doi: 10.1038/nrm2180 PubMedCrossRefGoogle Scholar
  5. Blower MD, Nachury M, Heald R, Weis K (2005) A Rae1-containing ribonucleoprotein complex is required for mitotic spindle assembly. Cell 121(2):223–234. doi: 10.1016/j.cell.2005.02.016 PubMedCrossRefGoogle Scholar
  6. Bono F, Gehring NH (2011) Assembly, disassembly and recycling: the dynamics of exon junction complexes. RNA Biol 8(1):24–29PubMedCrossRefGoogle Scholar
  7. Chichinadze K, Lazarashvili A, Tkemaladze J (2012) RNA in centrosomes: structure and possible functions. Protoplasma. doi: 10.1007/s00709-012-0422-6 PubMedGoogle Scholar
  8. Chopra P, Sethi G, Dastidar SG, Ray A (2010) Polo-like kinase inhibitors: an emerging opportunity for cancer therapeutics. Expert Opin Investig Drugs 19(1):27–43. doi: 10.1517/13543780903483191 PubMedCrossRefGoogle Scholar
  9. Filippova N, Yang X, King P, Nabors LB (2012) Phosphoregulation of the RNA-binding protein Hu antigen R (HuR) by Cdk5 affects centrosome function. J Biol Chem 287(38):32277–32287. doi: 10.1074/jbc.M112.353912 PubMedCrossRefGoogle Scholar
  10. Gajadhar A, Guha A (2010) A proximity ligation assay using transiently transfected, epitope-tagged proteins: application for in situ detection of dimerized receptor tyrosine kinases. Biotechniques 48(2):145–152. doi: 10.2144/000113354 PubMedCrossRefGoogle Scholar
  11. Gehring NH, Lamprinaki S, Kulozik AE, Hentze MW (2009) Disassembly of exon junction complexes by PYM. Cell 137(3):536–548. doi: 10.1016/j.cell.2009.02.042 PubMedCrossRefGoogle Scholar
  12. Groisman I, Huang YS, Mendez R, Cao Q, Theurkauf W, Richter JD (2000) CPEB, maskin, and cyclin B1 mRNA at the mitotic apparatus: implications for local translational control of cell division. Cell 103(3):435–447PubMedCrossRefGoogle Scholar
  13. Heidemann SR, Sander G, Kirschner MW (1977) Evidence for a functional role of RNA in centrioles. Cell 10(3):337–350PubMedCrossRefGoogle Scholar
  14. Inaki M, Kato D, Utsugi T, Onoda F, Hanaoka F, Murakami Y (2011) Genetic analyses using a mouse cell cycle mutant identifies magoh as a novel gene involved in Cdk regulation. Genes Cells 16(2):166–178. doi: 10.1111/j.1365-2443.2010.01479.x PubMedCrossRefGoogle Scholar
  15. Ishigaki Y, Nakamura Y, Takehara T, Shimasaki T, Tatsuno T, Takano F, Ueda Y, Motoo Y, Takegami T, Nakagawa H, Kuwabata S, Nemoto N, Tomosugi N, Miyazawa S (2011) Scanning electron microscopy with an ionic liquid reveals the loss of mitotic protrusions of cells during the epithelial-mesenchymal transition. Microsc Res Tech 74(11):1024–1031. doi: 10.1002/jemt.20989 PubMedCrossRefGoogle Scholar
  16. Ishigaki Y, Nakamura Y, Tatsuno T, Hashimoto M, Shimasaki T, Iwabuchi K, Tomosugi N (in press) Depletion of RNA-binding protein RBM8A (Y14) causes cell cycle deficiency and apoptosis in human cells. Exp Biol MedGoogle Scholar
  17. Ji JH, Jang YJ (2006) Screening of domain-specific target proteins of polo-like kinase 1: construction and application of centrosome/kinetochore-specific targeting peptide. J Biochem Mol Biol 39(6):709–716PubMedCrossRefGoogle Scholar
  18. Kataoka N, Diem MD, Kim VN, Yong J, Dreyfuss G (2001) Magoh, a human homolog of Drosophila mago nashi protein, is a component of the splicing-dependent exon–exon junction complex. EMBO J 20(22):6424–6433. doi: 10.1093/emboj/20.22.6424 PubMedCrossRefGoogle Scholar
  19. Kataoka N, Diem MD, Yoshida M, Hatai C, Dobashi I, Dreyfuss G, Hagiwara M, Ohno M (2011) Specific Y14 domains mediate its nucleo-cytoplasmic shuttling and association with spliced mRNA. Sci Rep 1:92. doi: 10.1038/srep00092 PubMedCentralPubMedCrossRefGoogle Scholar
  20. Kim TJ, Choi JJ, Kim WY, Choi CH, Lee JW, Bae DS, Son DS, Kim J, Park BK, Ahn G, Cho EY, Kim BG (2008) Gene expression profiling for the prediction of lymph node metastasis in patients with cervical cancer. Cancer Sci 99(1):31–38. doi: 10.1111/j.1349-7006.2007.00652.x PubMedGoogle Scholar
  21. Korzeniewski N, Hohenfellner M, Duensing S (2012) The centrosome as potential target for cancer therapy and prevention. Expert Opin Ther Targets. doi: 10.1517/14728222.2013.731396 PubMedGoogle Scholar
  22. Lambert JD, Nagy LM (2002) Asymmetric inheritance of centrosomally localized mRNAs during embryonic cleavages. Nature 420(6916):682–686. doi: 10.1038/nature01241 PubMedCrossRefGoogle Scholar
  23. Lau CK, Diem MD, Dreyfuss G, Van Duyne GD (2003) Structure of the Y14-Magoh core of the exon junction complex. Curr Biol 13(11):933–941PubMedCrossRefGoogle Scholar
  24. Le Hir H, Seraphin B (2008) EJCs at the heart of translational control. Cell 133(2):213–216. doi: 10.1016/j.cell.2008.04.002 PubMedCrossRefGoogle Scholar
  25. Le Hir H, Gatfield D, Braun IC, Forler D, Izaurralde E (2001a) The protein Mago provides a link between splicing and mRNA localization. EMBO Rep 2(12):1119–1124. doi: 10.1093/embo-reports/kve245 PubMedCentralPubMedCrossRefGoogle Scholar
  26. Le Hir H, Gatfield D, Izaurralde E, Moore MJ (2001b) The exon–exon junction complex provides a binding platform for factors involved in mRNA export and nonsense-mediated mRNA decay. EMBO J 20(17):4987–4997. doi: 10.1093/emboj/20.17.4987 PubMedCrossRefGoogle Scholar
  27. Lecuyer E, Yoshida H, Parthasarathy N, Alm C, Babak T, Cerovina T, Hughes TR, Tomancak P, Krause HM (2007) Global analysis of mRNA localization reveals a prominent role in organizing cellular architecture and function. Cell 131(1):174–187. doi: 10.1016/j.cell.2007.08.003 PubMedCrossRefGoogle Scholar
  28. Lee K, Rhee K (2011) PLK1 phosphorylation of pericentrin initiates centrosome maturation at the onset of mitosis. J Cell Biol 195(7):1093–1101. doi: 10.1083/jcb.201106093 PubMedCrossRefGoogle Scholar
  29. Ma HT, Poon RY (2011) How protein kinases co-ordinate mitosis in animal cells. Biochem J 435(1):17–31. doi: 10.1042/bj20100284 PubMedCrossRefGoogle Scholar
  30. Mahen R, Venkitaraman AR (2012) Pattern formation in centrosome assembly. Curr Opin Cell Biol 24(1):14–23. doi: 10.1016/j.ceb.2011.12.012 PubMedCrossRefGoogle Scholar
  31. Melero R, Buchwald G, Castano R, Raabe M, Gil D, Lazaro M, Urlaub H, Conti E, Llorca O (2012) The cryo-EM structure of the UPF–EJC complex shows UPF1 poised toward the RNA 3′ end. Nat Struct Mol Biol 19(5):498–505, S491–S492. doi: 10.1038/nsmb.2287 Google Scholar
  32. Mountzios G, Terpos E, Dimopoulos MA (2008) Aurora kinases as targets for cancer therapy. Cancer Treat Rev 34(2):175–182. doi: 10.1016/j.ctrv.2007.09.005 PubMedCrossRefGoogle Scholar
  33. Muromoto R, Taira N, Ikeda O, Shiga K, Kamitani S, Togi S, Kawakami S, Sekine Y, Nanbo A, Oritani K, Matsuda T (2009) The exon-junction complex proteins, Y14 and MAGOH regulate STAT3 activation. Biochem Biophys Res Commun 382(1):63–68. doi: 10.1016/j.bbrc.2009.02.127 PubMedCrossRefGoogle Scholar
  34. Ohbayashi N, Taira N, Kawakami S, Togi S, Sato N, Ikeda O, Kamitani S, Muromoto R, Sekine Y, Matsuda T (2008) An RNA biding protein, Y14 interacts with and modulates STAT3 activation. Biochem Biophys Res Commun 372(3):475–479. doi: 10.1016/j.bbrc.2008.05.073 PubMedCrossRefGoogle Scholar
  35. Preston JN, Trivedi MV (2012) Eribulin: a novel cytotoxic chemotherapy agent. Ann Pharmacother 46(6):802–811. doi: 10.1345/aph.1Q636 PubMedCrossRefGoogle Scholar
  36. Rabinowitz JS, Lambert JD (2010) Spiralian quartet developmental potential is regulated by specific localization elements that mediate asymmetric RNA segregation. Development 137(23):4039–4049. doi: 10.1242/dev.055269 PubMedCrossRefGoogle Scholar
  37. Salicioni AM, Xi M, Vanderveer LA, Balsara B, Testa JR, Dunbrack RL Jr, Godwin AK (2000) Identification and structural analysis of human RBM8A and RBM8B: two highly conserved RNA-binding motif proteins that interact with OVCA1, a candidate tumor suppressor. Genomics 69(1):54–62. doi: 10.1006/geno.2000.6315 PubMedCrossRefGoogle Scholar
  38. Sanhaji M, Friel CT, Wordeman L, Louwen F, Yuan J (2011) Mitotic centromere-associated kinesin (MCAK): a potential cancer drug target. Oncotarget 2(12):935–947PubMedGoogle Scholar
  39. Schmidt U, Richter K, Berger AB, Lichter P (2006) In vivo BiFC analysis of Y14 and NXF1 mRNA export complexes: preferential localization within and around SC35 domains. J Cell Biol 172(3):373–381. doi: 10.1083/jcb.200503061 PubMedCrossRefGoogle Scholar
  40. Schoffski P, Awada A, Dumez H, Gil T, Bartholomeus S, Wolter P, Taton M, Fritsch H, Glomb P, Munzert G (2012) A phase I, dose-escalation study of the novel Polo-like kinase inhibitor volasertib (BI 6727) in patients with advanced solid tumours. Eur J Cancer 48(2):179–186. doi: 10.1016/j.ejca.2011.11.001 PubMedCrossRefGoogle Scholar
  41. Silver DL, Watkins-Chow DE, Schreck KC, Pierfelice TJ, Larson DM, Burnetti AJ, Liaw HJ, Myung K, Walsh CA, Gaiano N, Pavan WJ (2010) The exon junction complex component Magoh controls brain size by regulating neural stem cell division. Nat Neurosci 13(5):551–558. doi: 10.1038/nn.2527 PubMedCentralPubMedCrossRefGoogle Scholar
  42. Singh G, Lykke-Andersen J (2003) New insights into the formation of active nonsense-mediated decay complexes. Trends Biochem Sci 28(9):464–466. doi: 10.1016/s0968-0004(03)00176-2 PubMedCrossRefGoogle Scholar
  43. Soderberg O, Gullberg M, Jarvius M, Ridderstrale K, Leuchowius KJ, Jarvius J, Wester K, Hydbring P, Bahram F, Larsson LG, Landegren U (2006) Direct observation of individual endogenous protein complexes in situ by proximity ligation. Nat Methods 3(12):995–1000. doi: 10.1038/nmeth947 PubMedCrossRefGoogle Scholar
  44. Sudo H, Tsuji AB, Sugyo A, Kohda M, Sogawa C, Yoshida C, Harada YN, Hino O, Saga T (2010) Knockdown of COPA, identified by loss-of-function screen, induces apoptosis and suppresses tumor growth in mesothelioma mouse model. Genomics 95(4):210–216. doi: 10.1016/j.ygeno.2010.02.002 PubMedCrossRefGoogle Scholar
  45. Wiese C, Zheng Y (1999) Gamma-tubulin complexes and their interaction with microtubule-organizing centers. Curr Opin Struct Biol 9(2):250–259. doi: 10.1016/s0959-440x(99)80035-9 PubMedCrossRefGoogle Scholar
  46. Zhao XF, Colaizzo-Anas T, Nowak NJ, Shows TB, Elliott RW, Aplan PD (1998) The mammalian homologue of mago nashi encodes a serum-inducible protein. Genomics 47(2):319–322. doi: 10.1006/geno.1997.5126 PubMedCrossRefGoogle Scholar
  47. Zhao XF, Nowak NJ, Shows TB, Aplan PD (2000) MAGOH interacts with a novel RNA-binding protein. Genomics 63(1):145–148. doi: 10.1006/geno.1999.6064 PubMedCrossRefGoogle Scholar
  48. Zheng Y, Jung MK, Oakley BR (1991) Gamma-tubulin is present in Drosophila melanogaster and Homo sapiens and is associated with the centrosome. Cell 65(5):817–823PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Yasuhito Ishigaki
    • 1
    Email author
  • Yuka Nakamura
    • 1
  • Takanori Tatsuno
    • 1
  • Mitsumasa Hashimoto
    • 2
    • 3
  • Kuniyoshi Iwabuchi
    • 3
  • Naohisa Tomosugi
    • 1
    • 4
    Email author
  1. 1.Medical Research InstituteKanazawa Medical UniversityKahokuJapan
  2. 2.Department of PhysicsKanazawa Medical UniversityKahokuJapan
  3. 3.Department of BiochemistryKanazawa Medical UniversityKahokuJapan
  4. 4.Medical Care Proteomics Biotechnology Co., Ltd.KahokuJapan

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