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Effect of hnRNP-like protein THO4 on growth and mRNA export in fission yeast

분열효모에서 hnRNP-유사 단백질인 THO4가 생장 및 mRNA 방출에 미치는 영향

  • Park, Jin Hee (School of Biological Science and Chemistry, Sungshin Women's University) ;
  • Lee, Sojeong (School of Biological Science and Chemistry, Sungshin Women's University) ;
  • Yoon, Jin Ho (School of Biological Science and Chemistry, Sungshin Women's University)
  • 박진희 (성신여자대학교 생명과학.화학부) ;
  • 이소정 (성신여자대학교 생명과학.화학부) ;
  • 윤진호 (성신여자대학교 생명과학.화학부)
  • Received : 2018.05.25
  • Accepted : 2018.06.11
  • Published : 2018.06.30

Abstract

The evolutionally conserved TREX complex member, Yra1/ALY, belongs to the REF (RNA and export factor binding proteins) family of hnRNP-like proteins, which has been implicated in multiple processes including transcription, nuclear RNA stability, and mRNA export. Fission yeast, Schizosaccharomyces pombe, genome encodes two members of REF proteins. In addition to Mlo3 known previously as an mRNA export factor, there is the other REF protein, Tho4, which is predicted as a component of THO complex. Here we showed that deletion of tho4 (SPBC106.12c) gene does not inhibit both growth and nuclear mRNA export. However, overexpression of tho4 displays growth retardation and slight accumulation of $poly(A)^+$ RNA in the nucleus. Neither ${\Delta}tho4$ ${\Delta}mlo3$ nor ${\Delta}tho4$ ${\Delta}mex67$ double mutants exhibit additive growth defect. Moreover, yeast two-hybrid and co-immunoprecipitation analysis did not show that the Tho4 protein interacted with any members of TREX complex and mRNA export factor Rae1. Contrary to expectation, these observations support that the S. pombe Tho4 is not a component of TREX complex, and not directly involved in bulk mRNA export from the nucleus.

진화적으로 보존된 TREX 복합체의 구성요소인 Yra1/ALY는 hnRNP-유사 단백질의 REF (RNA와 방출인자 결합단백질) 계열에 속하는 단백질로서 전사, 핵 RNA의 안정성, mRNA의 핵에서 방출 등 여러 과정에 관여하는 것으로 알려져 있다. 분열효모 Schizosaccharomyces pombe의 게놈에는 2개의 REF 단백질을 암호화하고 있다. 이미 mRNA 방출인자로 알려진 Mlo3 이외에 THO 복합체의 구성인자로 예측되는 Tho4이 존재한다. 본 연구에서 tho4 (SPBC106.12c) 유전자의 결실이 생장과 mRNA의 방출에 영향을 주지 않는다는 것을 알았다. 그러나 tho4 유전자를 과발현시키면 생장이 늦어지고 $poly(A)^+$ RNA가 핵 안에 약간 축적되었다. ${\Delta}tho4$ ${\Delta}mlo3$${\Delta}tho4$ ${\Delta}mex67$ 이 중 돌연변이는 어느 것도 추가적인 생장 결함을 보이지 않았다. 또한 Yeast two-hybrid와 공동침전(Co-immunoprecipitation) 분석에서 Tho4는 THO/TREX 복합체의 다른 구성인자들과 어떠한 상호작용도 보이지 않았다. 이와 같은 관찰결과들은 S. pombe의 Tho4 단백질이 기대와는 다르게THO/TREX 복합체의 구성인자가 아니며, mRNA 방출에도 직접 관여하지 않음을 의미한다.

Keywords

References

  1. Aguilera, A. 2002. The connection between transcription and genomic instability. EMBO J. 21, 195-201. https://doi.org/10.1093/emboj/21.3.195
  2. Alfa, C., Fantes, P., Hyams, J., Mcleod, M., and Warbrick, E. 1993. Experiments with fission yeast. Cold Spring Harbor, Cold Spring Harbor Laboratory, NY, USA.
  3. Bruhn, L., Munnerlyn, A., and Grosschedl, R. 1997. ALY, a context-dependent coactivator of LEF-1 and AML-1, is required for TCRalpha enhancer function. Genes Dev. 11, 640-653. https://doi.org/10.1101/gad.11.5.640
  4. Carmody, S.R. and Wente, S.R. 2009. mRNA nuclear export at a glance. J. Cell Sci. 122, 1933-1937. https://doi.org/10.1242/jcs.041236
  5. Chavez, S. and Aguilera, A. 1997. The yeast HPR1 gene has a functional role in transcriptional elongation that uncovers a novel source of genome instability. Genes Dev. 11, 3459-3470. https://doi.org/10.1101/gad.11.24.3459
  6. Cheng, H., Dufu, K., Lee, C.S., Hsu, J.L., Dias, A., and Reed, R. 2006. Human mRNA export machinery recruited to the 5' end of mRNA. Cell 127, 1389-1400. https://doi.org/10.1016/j.cell.2006.10.044
  7. Dominguez-Sanchez, M.S., Barroso, S., Gomez-Gonzalez, B., Luna, R., and Aguilera, A. 2011. Genome instability and transcription elongation impairment in human cells depleted of THO/TREX. PLoS Genet. 7, e1002386. https://doi.org/10.1371/journal.pgen.1002386
  8. Forsburg, S.L. and Sherman, D.A. 1997. General purpose tagging vectors for fission yeast. Gene 191, 191-195. https://doi.org/10.1016/S0378-1119(97)00058-9
  9. Gatfield, D. and Izaurralde, E. 2002. REF1/Aly and the additional exon junction complex proteins are dispensable for nuclear mRNA export. J. Cell Biol. 159, 579-588. https://doi.org/10.1083/jcb.200207128
  10. Heath, C.G., Viphakone, N., and Wilson, S.A. 2016. The role of TREX in gene expression and disease. Biochem. J. 473, 2911-2935. https://doi.org/10.1042/BCJ20160010
  11. Johnson, S.A., Kim, H., Erickson, B., and Bentley, D.L. 2011. The export factor Yra1 modulates mRNA 3' end processing. Nat. Struct. Mol. Biol. 18, 1164-1171. https://doi.org/10.1038/nsmb.2126
  12. Katahira, J. 2015. Nuclear export of messenger RNA. Gene 6, 163-184. https://doi.org/10.3390/genes6020163
  13. Koh, E. and Yoon, J.H. 2014. Effects of spThoc7 deletion on growth and mRNA export in fission yeast. Korean J. Microbiol. 50, 249-253. https://doi.org/10.7845/kjm.2014.4055
  14. Koh, E. and Yoon, J.H. 2015. Effects of Tho2, a component of THO complex, on growth and mRNA export in fission yeast. Korean J. Microbiol. 51, 181-185. https://doi.org/10.7845/kjm.2015.5020
  15. Lee, H. and Yoon, J.H. 2012. Effects of the repression of sphpr1 expression on growth and mRNA export in fission yeast. Korean J. Microbiol. 48, 171-174. https://doi.org/10.7845/kjm.2012.48.2.171
  16. Longman, D., Johnstone, I.L., and Caceres, J.F. 2003. The Ref/Aly proteins are dispensable for mRNA export and development in Caenorhabditis elegans. RNA 9, 881-891. https://doi.org/10.1261/rna.5420503
  17. Luna, R., Rondon, A.G., and Aguilera, A. 2012. New clues to understand the role of THO and other functionally related factors in mRNP biogenesis. Biochim. Biophys. Acta 1819, 514-520. https://doi.org/10.1016/j.bbagrm.2011.11.012
  18. Masuda, S., Das, R., Cheng, H., Hurt, E., Dorman, N., and Reed, R. 2005. Recruitment of the human TREX complex to mRNA during splicing. Genes Dev. 19, 1512-1517. https://doi.org/10.1101/gad.1302205
  19. Maundrell, K. 1993. Thiamine-repressible expression vectors pREP and pRIP for fission yeast. Gene 123, 127-130. https://doi.org/10.1016/0378-1119(93)90551-D
  20. Moreno, S., Klar, A., and Nurse, P. 1991. Molecular genetic analysis of fission yeast Schizosaccharomyces pombe. Methods Enzymol. 194, 795-823.
  21. Rodriguez-Navarro, S. and Hurt, E. 2011. Linking gene regulation to mRNA production and export. Curr. Opin. Cell Biol. 23, 302-309. https://doi.org/10.1016/j.ceb.2010.12.002
  22. Stewart, M. 2010. Nuclear export of mRNA. Trends Biochem. Sci. 35, 609-617. https://doi.org/10.1016/j.tibs.2010.07.001
  23. Strasser, K. and Hurt, E. 2000. Yra1p, a conserved nuclear RNA-binding protein, interacts directly with Mex67p and is required for mRNA export. EMBO J. 19, 410-420. https://doi.org/10.1093/emboj/19.3.410
  24. Strasser, K., Masuda, S., Mason, P., Pfannstiel, J., Oppizzi, M., Rodriguez-Navarro, S., Rondon, A.G., Aguilera, A., Struhl, K., Reed, R., et al. 2002. TREX is a conserved complex coupling transcription with messenger RNA export. Nature 417, 304-308. https://doi.org/10.1038/nature746
  25. Stutz, F., Bachi, A., Doerks, T., Braun, I.C., Seraphin, B., Wilm, M., Bork, P., and Izaurralde, E. 2000. REF, an evolutionary conserved family of hnRNP-like proteins, interacts with TAP/Mex67p and participates in mRNA nuclear export. RNA 6, 638-650. https://doi.org/10.1017/S1355838200000078
  26. Thakurta, A.G., Gopal, G., Yoon, J.H., Kozak, L., and Dhar, R. 2005. Homolog of BRCA2-interacting Dss1p and Uap56p link Mlo3p and Rae1p for mRNA export in fission yeast. EMBO J. 24, 2512-2523. https://doi.org/10.1038/sj.emboj.7600713
  27. Walsh, M.J., Hautbergue, G.M., and Wilson, S.A. 2010. Structure and function of mRNA export adaptors. Biochem. Soc. Trans. 38, 232-236. https://doi.org/10.1042/BST0380232
  28. Yoh, S.M., Cho, H., Pickle, L., Evans, R.M., and Jones, K.A. 2007. The Spt6 SH2 domain binds Ser2-P RNAPII to direct Iws1-dependent mRNA splicing and export. Genes Dev. 21, 160-174. https://doi.org/10.1101/gad.1503107
  29. Yoon, J.H., Love, D., Guhathakurta, A., Hanover, J.A., and Dhar, R. 2000. Mex67p of Schizosaccharomyces pombe interacts with Rae1p in mediating mRNA export. Mol. Cell. Biol. 20, 8767-8782. https://doi.org/10.1128/MCB.20.23.8767-8782.2000
  30. Zenklusen, D., Vinciguerra, P., Strahm, Y., and Stutz, F. 2001. The yeast hnRNP-like proteins Yra1p and Yra2p participate in mRNA export through interaction with Mex67p. Mol. Cell. Biol. 21, 4219-4232. https://doi.org/10.1128/MCB.21.13.4219-4232.2001