Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
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Identification and analysis of microRNAs in Candida albicans

Candida albicans의 마이크로RNA 동정과 분석

  • Cho, Jin-Hyun (Department of Prosthodontics, Kyungpook National University, School of Dentistry) ;
  • Lee, Heon-Jin (Department of Microbiology and Immunology, Kyungpook National University, School of Dentistry)
  • 조진현 (경북대학교 치과대학 보철학교실) ;
  • 이헌진 (경북대학교 치과대학 구강미생물학교실)
  • Received : 2017.09.21
  • Accepted : 2017.11.15
  • Published : 2017.12.30
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Abstract

Oral infection due to Candida albicans is a widely recognized and frequent cause of superficial infections of the oral mucosa (oral candidiasis). Although oral candidiasis is not a life-threatening fungemia, it can cause severe problems in individuals under certain conditions. MicroRNAs (miRNAs) are noncoding, small RNA molecules, which regulate the expression of other genes by inhibiting the translation of target mRNAs. The present study was designed to identify miRNAs in C. albicans and determine their possible roles in this organism. miRNA-sized small RNAs (msRNAs) were cloned in C. albicans by deep sequencing, and their secondary structures were analyzed. All the cloned msRNAs satisfied conditions required to qualify them as miRNAs. Bioinformatics analysis revealed that two of the most highly expressed C. albicans msRNAs, Ca-363 and Ca-2019, were located in the 3' untranslated region of the corticosteroid-binding protein 1 (CBP1) gene in a reverse orientation. miRNA mimics were transformed into C. albicans to investigate their RNA-inhibitory functions. RNA oligonucleotide-transformed C. albicans was then observed by fluorescent microscopy. Quantitative PCR analysis showed that these msRNAs did not inhibit CBP1 gene expression 4 hr and 8 hr after ectopic miRNA transformation. These results suggest that msRNAs in C. albicans possess an miRNA-triggered RNA interference gene-silencing function, which is distinct from that exhibited by other eukaryotic systems.

Candida albicans에 의한 구강 감염(캔디다증)은 구강 점막에 빈번하게 발생하며 잘 알려진 질병이다. 구강 캔디다증은 생명을 위협하는 정도의 곰팡이 감염증은 아니나, 특정상황에서 개인에게 심각한 위험을 초래할 수도 있다. 마이크로 RNA는 세포 내에서 다른 타겟 유전자를 저해하는 작은 크기의 RNA 분자이며 단백질을 코딩하지는 않고 번역과정을 억제하는 조절자로서의 역할을 하고 있다. 본 연구는 C. albicans의 마이크로RNA를 처음으로 동정하고 그러한 마이크로RNA가 지닌 기능을 조사하기 위함이다. 이를 위하여 C. albicans의 small RNA를 차세대 염기분석법을 통하여 분석하고 그러한 RNA들의 2차 구조를 생물정보학적 방법으로 조사하였다. 분석한 small RNA들은 마이크로 RNA라고 불리울 수 있는 특징들을 가지고 있었으며, 특별히 높게 발현되고 있는 두개의 마이크로 RNA 정도 크기의 RNA가 CBP1 유전자의 3' 말단 비번역구역(UTR)에서 반대방향으로 발현하는 것을 밝혀 내었다. 우리는 이러한 C. albicans의 RNA가 CBP1 유전자를 타겟으로 하여 조절하는지 알아보기 위해 RNA를 인위적으로 합성한 후 세포 내로 주입하고, 형광형미경으로 도입 사실을 확인하였다. 하지만 4시간과 8시간 후에 CBP1의 발현 변화는 관찰되지 않았다. 따라서, 이러한 결과는 C. albicans가 마이크로RNA에 의한 RNA 간섭(RNAi) 작용이 다른 진핵세포와는 다르게 작용하는 것을 알 수 있다.

Keywords

References

  1. Ambros, V., Bartel, B., Bartel D. P., Burge, C. B., Carrington, J. C., Chen, X., Dreyfuss, G., Eddy, S. R., Griffiths-Jones, S., Marchall, M., Matzke, M., Ruvkun, G. and Tuschl, T. 2003. A uniform system for microRNA annotation. RNA 9, 277-279. https://doi.org/10.1261/rna.2183803
  2. Bernstein, D. A., Vyas, V. K., Weinberg, D. E., Drinnenberg, I. A., Bartel, D. P. and Fink, G. R. 2012. Candida albicans Dicer (CaDcr1) is required for efficient ribosomal and spliceosomal RNA maturation. Proc. Natl. Acad. Sci. USA 109, 523-528. https://doi.org/10.1073/pnas.1118859109
  3. Celluzzi, A. and Masotti, A. 2016. How our other genome controls our epi-genome. Trends. Microbiol. 24, 1-11. https://doi.org/10.1016/j.tim.2015.10.010
  4. Choi, J. W., Um, J. H., Cho, J. H. and Lee, H. J. 2017. Tiny RNAs and their voyage via extracellular vesicles: Secretion of bacterial small RNA and eukaryotic microRNA. Exp. Biol. Med. 242, 1475-1481. https://doi.org/10.1177/1535370217723166
  5. Drinnenberg, I. A., Weinberg, D. E., Xie, K. T., Mower, J. P., Wolfe, K. H., Fink, G. R. and Bartel, D. P. 2009. RNAi in budding yeast. Science 326, 544-550. https://doi.org/10.1126/science.1176945
  6. Gil-Bona, A., Llama-Palacios, A., Parra, C. M., Vivanco, F., Nombela, C., Monteoliva, L. and Gil, C. 2015. Proteomics unravels extracellular vesicles as carriers of classical cytoplasmic proteins in Candida albicans. J. Proteome. Res. 14, 142-153. https://doi.org/10.1021/pr5007944
  7. Hafner, M., Landgraf, P., Ludwig, J., Rice, A., Ojo, T., Lin, C., Holoch, D., Lim, C. and Tuschl, T. 2008. Identification of microRNAs and other small regulatory RNAs using cDNA library sequencing. Methods 4, 3-12.
  8. Lee, H. J. 2013. Exceptional stories of microRNAs. Exp. Biol. Med. 238, 339-343. https://doi.org/10.1258/ebm.2012.012251
  9. Lee, H. J. 2014. Additional stories of microRNAs. Exp. Biol. Med. 239, 1275-1279. https://doi.org/10.1177/1535370214544269
  10. Lee, H. J. and Hong, S. H. 2012. Analysis of microRNA-size, small RNAs in Streptococcus mutans by deep sequencing. FEMS Microbiol. Lett. 326, 131-136. https://doi.org/10.1111/j.1574-6968.2011.02441.x
  11. Nobile, C. J and Johnson, A. D. 2015. Candida albicans Biofilms and Human Disease. Annu. Rev. Microbiol. 69, 71-92. https://doi.org/10.1146/annurev-micro-091014-104330
  12. Okamura, K., Phillips, M. D., Tyler, D. M., Duan, H., Chou, Y. T. and Lai, E. C. 2008. The regulatory activity of microRNA* species has substantial influence on microRNA and 3' UTR evolution. Nat. Struct. Mol. Biol. 15, 354-363. https://doi.org/10.1038/nsmb.1409
  13. O'Toole, A. S., Miller, S., Haines, N., Zink, M. C. and Serra, M. J. 2006. Comprehensive thermodynamic analysis of 3' double-nucleotide overhangs neighboring Watson-Crick terminal base pairs. Nucleic Acids Res. 34, 3338-3344. https://doi.org/10.1093/nar/gkl428
  14. Rodrigues, M. L., Nimrichter, L., Oliveira, D. L., Nosanchuk, J. D. and Casadevall, A. 2008. Vesicular trans-cell wall transport in fungi: a mechanism for the delivery of virulence-associated macromolecules? Lipid Insights 2, 27-40.
  15. Salvatori, O., Puri, S., Tati, S. and Edgerton, M. 2016. Innate immunity and saliva in Candida albicans-mediated oral diseases. J. Dent. Res. 95, 365-371. https://doi.org/10.1177/0022034515625222
  16. Staab, J. F., White, T. C. and Marr, K. A. 2010. Hairpin dsRNA does not trigger RNA interference in Candida al cans cells. Yeast 28, 1-8.
  17. Vargas, G., Rocha, J. D. B., Oliveira, D. L., Albuquerque, P. C., Frases, S., Santos, S. S., Nosanchuk, J. D., Gomes, A. M. O., Medeiros, L. C. A. S., Miranda, K., Sobreira, T. J. P., Nakayasu, E. S., Arigi, E. A., Casadevall, A., Guimaraes, A. J., Rodrigues, M. L., Freire-de-Lima, C. G., Almeida, I. C and Nimrichter, L. 2015. Compositional and immunobiological analyses of extracellular vesicles released by Candida albicans. Cell Microbiol. 17, 389-407. https://doi.org/10.1111/cmi.12374
  18. Zuker, M. 2003. Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res. 31, 3406-3415. https://doi.org/10.1093/nar/gkg595