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Filamentous growth of Escherichia coli by dephosphorylated NPr

탈인산화된 NPr에 의한 대장균의 섬유상 생장

  • Choi, Umji (Department of Bioscience and Bioinformatics, Myongji University) ;
  • Seok, Yeong-Jae (Department of Biological Sciences and Institute of Microbiology, Seoul National University) ;
  • Lee, Chang-Ro (Department of Bioscience and Bioinformatics, Myongji University)
  • 최엄지 (명지대학교 생명과학정보학과) ;
  • 석영재 (서울대학교 생명과학부) ;
  • 이창로 (명지대학교 생명과학정보학과)
  • Received : 2017.05.02
  • Accepted : 2017.06.29
  • Published : 2017.09.30

Abstract

The nitrogen phosphotransferase (PTS) system is a regulatory cascade present in most Proteobacteria, where it controls different functions. The nitrogen PTS is usually composed of $EI^{Ntr}$ (encoded by the ptsP gene), NPr (encoded by the ptsO gene), and $EIIA^{Ntr}$ (encoded by the ptsN gene). While $EIIA^{Ntr}$ plays a role in a variety of cellular processes, such as potassium homeostasis, regulation of ppGpp accumulation, nitrogen and carbon metabolisms, and regulation of ABC transporters, little information is available for a physiological role of NPr. A recent study showed that dephosphorylated NPr affects adaptation to envelope stresses in Escherichia coli. In this study, we provide another phenotype related to NPr. The ptsP mutant showed a filamentation phenotype. The filamentation phenotype of the ptsP mutant was recovered by additional deletion of the ptsO gene, but not by additional deletion of the ptsN gene, suggesting that an increased level of dephosphorylated NPr in the ptsP mutant renders cells the filamentous growth. This idea was confirmed by the fact that cells with increased levels of dephosphorylated NPr shows the filamentation phenotype. Additionally, we showed that cell size of E. coli increases with incremental dephosphorylated NPr concentrations. These results suggested that dephosphorylated NPr induces morphological change of E. coli.

대부분의 Proteobacteria에 존재하는 질소 인산전달계는 다양한 세포내 조절에 관여하는 cascade이다. 이들은 ptsP 유전자에 의해 암호화되는 $EI^{Ntr}$, ptsO에 의해 암호화되는 NPr, ptsN에 의해 암호화되는 $EIIA^{Ntr}$로 이루어져 있다. 이들 중 $EIIA^{Ntr}$$K^+$ 농도 조절, ppGpp 농도 조절, 질소와 탄소 대사, ABC transporter의 조절 등 다양한 세포내 조절과정에 관여하지만, NPr의 생리적 기능에 대해서는 알려진 바가 많지 않다. 최근의 한 논문은 대장균에서 탈인산화된 NPr이 세포막 스트레스 반응에 관여한다는 사실이 밝혔다. 본 연구에서는 NPr과 관련된 새로운 표현형을 제공한다. ptsP 유전자가 결손된 균주는 filamentation 표현형을 나타내었다. ptsP 결손균주의 이런 표현형은 ptsO 유전자의 추가적인 결실에 의해 사라졌지만, ptsN 유전자의 추가적 소실에 의해서는 유지되었다. 이는 ptsP 결손균주의 filamentation 표현형이 탈인산화된 NPr의 증가 때문에 나타났음을 나타낸다. 이런 생각은 야생종에서 탈인산화된 NPr이 증가되었을 때 filamentation 표현형을 나타낸다는 사실을 통해 확증되었다. 또한 탈인산화된 NPr의 양이 증가함에 따라 대장균의 세포 길이가 점진적으로 증가한다는 사실을 알 수 있었다. 이러한 결과는 탈인산화된 NPr이 대장균의 형태적 변화를 유도함을 시사한다.

Keywords

References

  1. Chen, J.C. and Beckwith, J. 2001. FtsQ, FtsL and FtsI require FtsK, but not FtsN, for co-localization with FtsZ during Escherichia coli cell division. Mol. Microbiol. 42, 395-413. https://doi.org/10.1046/j.1365-2958.2001.02640.x
  2. Choi, J., Shin, D., Yoon, H., Kim, J., Lee, C.R., Kim, M., Seok, Y.J., and Ryu, S. 2010. Salmonella pathogenicity island 2 expression negatively controlled by $EIIA^{Ntr}$-SsrB interaction is required for Salmonella virulence. Proc. Natl. Acad. Sci. USA 107, 20506-20511. https://doi.org/10.1073/pnas.1000759107
  3. Deutscher, J., Francke, C., and Postma, P.W. 2006. How phosphotransferase system-related protein phosphorylation regulates carbohydrate metabolism in bacteria. Microbiol. Mol. Biol. Rev. 70, 939-1031. https://doi.org/10.1128/MMBR.00024-06
  4. Dozot, M., Poncet, S., Nicolas, C., Copin, R., Bouraoui, H., Maze, A., Deutscher, J., De Bolle, X., and Letesson, J.J. 2010. Functional characterization of the incomplete phosphotransferase system (PTS) of the intracellular pathogen Brucella melitensis. PLoS One 5, e12679. https://doi.org/10.1371/journal.pone.0012679
  5. Gopel, Y., Papenfort, K., Reichenbach, B., Vogel, J., and Gorke, B. 2013. Targeted decay of a regulatory small RNA by an adaptor protein for RNase E and counteraction by an anti-adaptor RNA. Genes Dev. 27, 552-564. https://doi.org/10.1101/gad.210112.112
  6. Kaddor, C. and Steinbuchel, A. 2011. Effects of homologous phosphoenolpyruvate-carbohydrate phosphotransferase system proteins on carbohydrate uptake and poly(3-Hydroxybutyrate) accumulation in Ralstonia eutropha H16. Appl. Environ. Microbiol. 77, 3582-3590. https://doi.org/10.1128/AEM.00218-11
  7. Kalamorz, F., Reichenbach, B., Marz, W., Rak, B., and Gorke, B. 2007. Feedback control of glucosamine-6-phosphate synthase GlmS expression depends on the small RNA GlmZ and involves the novel protein YhbJ in Escherichia coli. Mol. Microbiol. 65, 1518-1533. https://doi.org/10.1111/j.1365-2958.2007.05888.x
  8. Karstens, K., Zschiedrich, C.P., Bowien, B., Stulke, J., and Gorke, B. 2014. Phosphotransferase protein $EIIA^{Ntr}$ interacts with SpoT, a key enzyme of the stringent response, in Ralstonia eutropha H16. Microbiology 160, 711-722. https://doi.org/10.1099/mic.0.075226-0
  9. Kim, H.J., Lee, C.R., Kim, M., Peterkofsky, A., and Seok, Y.J. 2011. Dephosphorylated NPr of the nitrogen PTS regulates lipid A biosynthesis by direct interaction with LpxD. Biochem. Biophys. Res. Commun. 409, 556-561. https://doi.org/10.1016/j.bbrc.2011.05.044
  10. King, N.D. and O'Brian, M.R. 2001. Evidence for direct interaction between enzyme $I^{Ntr}$ and aspartokinase to regulate bacterial oligopeptide transport. J. Biol. Chem. 276, 21311-21316. https://doi.org/10.1074/jbc.M101982200
  11. Koo, B.M., Yoon, M.J., Lee, C.R., Nam, T.W., Choe, Y.J., Jaffe, H., Peterkofsky, A., and Seok, Y.J. 2004. A novel fermentation/respiration switch protein regulated by enzyme $IIA^{Glc}$ in Escherichia coli. J. Biol. Chem. 279, 31613-31621. https://doi.org/10.1074/jbc.M405048200
  12. Luttmann, D., Gopel, Y., and Gorke, B. 2012. The phosphotransferase protein $EIIA^{Ntr}$ modulates the phosphate starvation response through interaction with histidine kinase PhoR in Escherichia coli. Mol. Microbiol. 86, 96-110. https://doi.org/10.1111/j.1365-2958.2012.08176.x
  13. Luttmann, D., Heermann, R., Zimmer, B., Hillmann, A., Rampp, I.S., Jung, K., and Gorke, B. 2009. Stimulation of the potassium sensor KdpD kinase activity by interaction with the phosphotransferase protein $IIA^{Ntr}$ in Escherichia coli. Mol. Microbiol. 72, 978-994. https://doi.org/10.1111/j.1365-2958.2009.06704.x
  14. Lee, S.J., Boos, W., Bouche, J.P., and Plumbridge, J. 2000. Signal transduction between a membrane-bound transporter, PtsG, and a soluble transcription factor, Mlc, of Escherichia coli. EMBO J. 19, 5353-5361. https://doi.org/10.1093/emboj/19.20.5353
  15. Lee, C.R., Cho, S.H., Yoon, M.J., Peterkofsky, A., and Seok, Y.J. 2007. Escherichia coli enzyme $IIA^{Ntr}$ regulates the $K^+$ transporter TrkA. Proc. Natl. Acad. Sci. USA 104, 4124-4129. https://doi.org/10.1073/pnas.0609897104
  16. Lee, K.J., Jeong, C.S., An, Y.J., Lee, H.J., Park, S.J., Seok, Y.J., Kim, P., Lee, J.H., Lee, K.H., and Cha, S.S. 2011. FrsA functions as a cofactor-independent decarboxylase to control metabolic flux. Nat. Chem. Biol. 7, 434-436. https://doi.org/10.1038/nchembio.589
  17. Lee, J., Park, Y.H., Kim, Y.R., Seok, Y.J., and Lee, C.R. 2015. Dephosphorylated NPr is involved in an envelope stress response of Escherichia coli. Microbiology 161, 1113-1123. https://doi.org/10.1099/mic.0.000056
  18. Nam, T.W., Cho, S.H., Shin, D., Kim, J.H., Jeong, J.Y., Lee, J.H., Roe, J.H., Peterkofsky, A., Kang, S.O., Ryu, S., et al. 2001. The Escherichia coli glucose transporter enzyme $IICB^{Glc}$ recruits the global repressor Mlc. EMBO J. 20, 491-498. https://doi.org/10.1093/emboj/20.3.491
  19. Park, Y.H., Lee, C.R., Choe, M., and Seok, Y.J. 2013. HPr antagonizes the $anti-s^{70}$ activity of Rsd in Escherichia coli. Proc. Natl. Acad. Sci. USA 110, 21142-21147. https://doi.org/10.1073/pnas.1316629111
  20. Park, Y.H., Lee, B.R., Seok, Y.J., and Peterkofsky, A. 2006. In vitro reconstitution of catabolite repression in Escherichia coli. J. Biol. Chem. 281, 6448-6454. https://doi.org/10.1074/jbc.M512672200
  21. Peterkofsky, A., Wang, G., and Seok, Y.J. 2006. Parallel PTS systems. Arch. Biochem. Biophys. 453, 101-107. https://doi.org/10.1016/j.abb.2006.01.004
  22. Pfluger-Grau, K. and Gorke, B. 2010. Regulatory roles of the bacterial nitrogen-related phosphotransferase system. Trends Microbiol. 18, 205-214. https://doi.org/10.1016/j.tim.2010.02.003
  23. Postma, P.W., Lengeler, J.W., and Jacobson, G.R. 1993. Phosphoenolpyruvate: carbohydrate phosphotransferase systems of bacteria. Microbiol. Rev. 57, 543-594.
  24. Prell, J., Mulley, G., Haufe, F., White, J.P., Williams, A., Karunakaran, R., Downie, J.A., and Poole, P.S. 2012. The $PTS^{Ntr}$ system globally regulates ATP-dependent transporters in Rhizobium leguminosarum. Mol. Microbiol. 84, 117-129. https://doi.org/10.1111/j.1365-2958.2012.08014.x
  25. Rabus, R., Reizer, J., Paulsen, I., and Saier, M.H.Jr. 1999. Enzyme $I^{Ntr}$ from Escherichia coli. A novel enzyme of the phosphoenolpyruvate-dependent phosphotransferase system exhibiting strict specificity for its phosphoryl acceptor, NPr. J. Biol. Chem. 274, 26185-26191. https://doi.org/10.1074/jbc.274.37.26185
  26. Rhodius, V.A., Suh, W.C., Nonaka, G., West, J., and Gross, C.A. 2006. Conserved and variable functions of the ${\sigma}^E$ stress response in related genomes. PLoS Biol. 4, e2.
  27. Ronneau, S., Petit, K., De Bolle, X., and Hallez, R. 2016. Phosphotransferase-dependent accumulation of (p)ppGpp in response to glutamine deprivation in Caulobacter crescentus. Nat. Commun. 7, 11423. https://doi.org/10.1038/ncomms11423
  28. Sperandeo, P., Cescutti, R., Villa, R., Di Benedetto, C., Candia, D., Deho, G., and Polissi, A. 2007. Characterization of lptA and lptB, two essential genes implicated in lipopolysaccharide transport to the outer membrane of Escherichia coli. J. Bacteriol. 189, 244-253. https://doi.org/10.1128/JB.01126-06
  29. Tanaka, Y., Kimata, K., and Aiba, H. 2000. A novel regulatory role of glucose transporter of Escherichia coli: membrane sequestration of a global repressor Mlc. EMBO J. 19, 5344-5352. https://doi.org/10.1093/emboj/19.20.5344
  30. Velazquez, F., Pflüger, K., Cases, I., De Eugenio, L.I., and de Lorenzo, V. 2007. The phosphotransferase system formed by PtsP, PtsO, and PtsN proteins controls production of polyhydroxyalkanoates in Pseudomonas putida. J. Bacteriol. 189, 4529-4533. https://doi.org/10.1128/JB.00033-07
  31. Wissel, M.C., Wendt, J.L., Mitchell, C.J., and Weiss, D.S. 2005. The transmembrane helix of the Escherichia coli division protein FtsI localizes to the septal ring. J. Bacteriol. 187, 320-328. https://doi.org/10.1128/JB.187.1.320-328.2005
  32. Yang, D.C., Peters, N.T., Parzych, K.R., Uehara, T., Markovski, M., and Bernhardt, T.G. 2011. An ATP-binding cassette transporter-like complex governs cell-wall hydrolysis at the bacterial cytokinetic ring. Proc. Natl. Acad. Sci. USA 108, E1052-1060. https://doi.org/10.1073/pnas.1107780108
  33. Yoo, W., Yoon, H., Seok, Y.J., Lee, C.R., Lee, H.H., and Ryu, S. 2016. Fine-tuning of amino sugar homeostasis by $EIIA^{Ntr}$ in Salmonella Typhimurium. Sci. Rep. 6, 33055. https://doi.org/10.1038/srep33055