Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

The metabolic enzyme CTP synthase forms cytoskeletal filaments



Filament-forming cytoskeletal proteins are essential for the structure and organization of all cells. Bacterial homologues of the major eukaryotic cytoskeletal families have now been discovered, but studies suggest that yet more remain to be identified. We demonstrate that the metabolic enzyme CTP synthase (CtpS) forms filaments in Caulobacter crescentus. CtpS is bifunctional, as the filaments it forms regulate the curvature of C. crescentus cells independently of its catalytic function. The morphogenic role of CtpS requires its functional interaction with the intermediate filament, crescentin (CreS). Interestingly, the Escherichia coli CtpS homologue also forms filaments both in vivo and in vitro, suggesting that CtpS polymerization may be widely conserved. E. coli CtpS can replace the enzymatic and morphogenic functions of C. crescentus CtpS, indicating that C. crescentus has adapted a conserved filament-forming protein for a secondary role. These results implicate CtpS as a novel bifunctional member of the bacterial cytoskeleton and suggest that localization and polymerization may be important properties of metabolic enzymes.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Rent or buy this article

Prices vary by article type



Prices may be subject to local taxes which are calculated during checkout

Figure 1: mCherry–CtpS is dynamic and co-localizes with linear filamentous structures along the inner curvature of C. crescentus cells.
Figure 2: CtpS forms filaments in C. crescentus.
Figure 3: CtpS regulates C. crescentus cell shape independently of its enzymatic activity.
Figure 4: CtpS regulates cell shape through an interaction with CreS.
Figure 5: The E. coli CtpS homologue forms filaments both in vivo and in vitro.


  1. Bi, E. F. & Lutkenhaus, J. FtsZ ring structure associated with division in Escherichia coli. Nature 354, 161–164 (1991).

    Article  CAS  Google Scholar 

  2. Lowe, J. & Amos, L. A. Crystal structure of the bacterial cell-division protein FtsZ. Nature 391, 203–206 (1998).

    Article  CAS  Google Scholar 

  3. Jones, L. J., Carballido-Lopez, R. & Errington, J. Control of cell shape in bacteria: helical, actin-like filaments in Bacillus subtilis. Cell 104, 913–922 (2001).

    Article  CAS  Google Scholar 

  4. Ausmees, N., Kuhn, J. R. & Jacobs-Wagner, C. The bacterial cytoskeleton: an intermediate filament-like function in cell shape. Cell 115, 705–713 (2003).

    Article  CAS  Google Scholar 

  5. Briegel, A. et al. Multiple large filament bundles observed in Caulobacter crescentus by electron cryotomography. Mol. Microbiol. 62, 5–14 (2006).

    Article  CAS  Google Scholar 

  6. Li, Z. & Jensen, G. J. Electron cryotomography: a new view into microbial ultrastructure. Curr. Opin. Microbiol. 12, 333–340 (2009).

    Article  CAS  Google Scholar 

  7. Li, Z., Trimble, M. J., Brun, Y. V. & Jensen, G. J. The structure of FtsZ filaments in vivo suggests a force-generating role in cell division. EMBO J. 26, 4694–4708 (2007).

    Article  CAS  Google Scholar 

  8. Werner, J. N. et al. Quantitative genome-scale analysis of protein localization in an asymmetric bacterium. Proc. Natl Acad. Sci. USA 106, 7858–7863 (2009).

    Article  CAS  Google Scholar 

  9. Long, C. W., Levitzki, A. & Koshland, D. E., Jr. The subunit structure and subunit interactions of cytidine triphosphate synthetase. J. Biol. Chem. 245, 80–87 (1970).

    CAS  PubMed  Google Scholar 

  10. Briegel, A. et al. Location and architecture of the Caulobacter crescentus chemoreceptor array. Mol. Microbiol. 69, 30–41 (2008).

    Article  CAS  Google Scholar 

  11. Ma, X., Ehrhardt, D. W. & Margolin, W. Colocalization of cell division proteins FtsZ and FtsA to cytoskeletal structures in living Escherichia coli cells by using green fluorescent protein. Proc. Natl Acad. Sci. USA 93, 12998–13003 (1996).

    Article  CAS  Google Scholar 

  12. Gitai, Z., Dye, N. & Shapiro, L. An actin-like gene can determine cell polarity in bacteria. Proc. Natl Acad. Sci. USA 101, 8643–8648 (2004).

    Article  CAS  Google Scholar 

  13. Hartman, S. C. The interaction of 6-diazo-5-oxo-L-norleucine with phosphoribosyl pyrophosphate amidotransferase. J. Biol. Chem. 238, 3036–3047 (1963).

    CAS  PubMed  Google Scholar 

  14. Karimova, G., Pidoux, J., Ullmann, A. & Ladant, D. A bacterial two-hybrid system based on a reconstituted signal transduction pathway. Proc. Natl Acad. Sci. USA 95, 5752–5756 (1998).

    Article  CAS  Google Scholar 

  15. Srinivasan, R., Mishra, M., Murata-Hori, M. & Balasubramanian, M. K. Filament formation of the Escherichia coli actin-related protein, MreB, in fission yeast. Curr. Biol. 17, 266–272 (2007).

    Article  CAS  Google Scholar 

  16. Srinivasan, R., Mishra, M., Wu, L., Yin, Z. & Balasubramanian, M. K. The bacterial cell division protein FtsZ assembles into cytoplasmic rings in fission yeast. Genes Dev. 22, 1741–1746 (2008).

    Article  CAS  Google Scholar 

  17. Levitzki, A. & Koshland, D. E., Jr. Cytidine triphosphate synthetase. Covalent intermediates and mechanisms of action. Biochemistry 10, 3365–3371 (1971).

    Article  CAS  Google Scholar 

  18. Endrizzi, J. A., Kim, H., Anderson, P. M. & Baldwin, E. P. Crystal structure of Escherichia coli cytidine triphosphate synthetase, a nucleotide-regulated glutamine amidotransferase/ATP-dependent amidoligase fusion protein and homologue of anticancer and antiparasitic drug targets. Biochemistry 43, 6447–6463 (2004).

    Article  CAS  Google Scholar 

  19. Lunn, F. A., Macleod, T. J. & Bearne, S. L. Mutational analysis of conserved glycine residues 142, 143 and 146 reveals Gly(142) is critical for tetramerization of CTP synthase from Escherichia coli. Biochem. J. 412, 113–121 (2008).

    Article  CAS  Google Scholar 

  20. Paluh, J. L., Zalkin, H., Betsch, D. & Weith, H. L. Study of anthranilate synthase function by replacement of cysteine 84 using site-directed mutagenesis. J. Biol. Chem. 260, 1889–1894 (1985).

    CAS  PubMed  Google Scholar 

  21. Charbon, G., Cabeen, M. T. & Jacobs-Wagner, C. Bacterial intermediate filaments: in vivo assembly, organization, and dynamics of crescentin. Genes Dev. 23, 1131–1144 (2009).

    Article  CAS  Google Scholar 

  22. Cabeen, M. T. et al. Bacterial cell curvature through mechanical control of cell growth. EMBO J. 28, 1208–1219 (2009).

    Article  CAS  Google Scholar 

  23. Kleinschmidt, A. K., Moss, J. & Lane, D. M. Acetyl coenzyme A carboxylase: filamentous nat of the animal enzymes. Science 166, 1276–1278 (1969).

    Article  CAS  Google Scholar 

  24. Carrey, E. A. et al. Detection and location of the enzymes of de novo pyrimidine biosynthesis in mammalian spermatozoa. Reproduction 123, 757–768 (2002).

    Article  CAS  Google Scholar 

  25. Zigler, J. S., Jr. & Rao, P. V. Enzyme/crystallins and extremely high pyridine nucleotide levels in the eye lens. FASEB J. 5, 223–225 (1991).

    Article  CAS  Google Scholar 

  26. Bork, P., Sander, C. & Valencia, A. An ATPase domain common to prokaryotic cell cycle proteins, sugar kinases, actin and hsp70 heat shock proteins. Proc. Natl Acad. Sci. USA 89, 7290–7294 (1992).

    Article  CAS  Google Scholar 

  27. Evinger, M. & Agabian, N. Envelope-associated nucleoid from Caulobacter crescentus stalked and swarmer cells. J. Bacteriol. 132, 294–301 (1977).

    CAS  PubMed  PubMed Central  Google Scholar 

  28. Pinter, S. F., Aubert, S. D. & Zakian, V. A. The Schizosaccharomyces pombe Pfh1p DNA helicase is essential for the maintenance of nuclear and mitochondrial DNA. Mol. Cell Biol. 28, 6594–6608 (2008).

    Article  CAS  Google Scholar 

  29. Ely, B. Genetics of Caulobacter crescentus. Methods Enzymol. 204, 372–384 (1991).

    Article  CAS  Google Scholar 

  30. Ellgaard, A. K., Mullin, D. A. and Minnich, S. A. Plasmid transformation of Caulobacter crescentus using electroporation. Abstr. Annu. Meeting Am. Soc. Microbiol. 209, 208 (1989).

    Google Scholar 

  31. Meisenzahl, A. C., Shapiro, L. & Jenal, U. Isolation and characterization of a xylose-dependent promoter from Caulobacter crescentus. J. Bacteriol. 179, 592–600 (1997).

    Article  CAS  Google Scholar 

  32. Iancu, C. V. et al. Electron cryotomography sample preparation using the Vitrobot. Nat. Protoc. 1, 2813–2819 (2007).

    Article  Google Scholar 

  33. Tivol, W., Briegel, A. & Jensen, G. J. An improved cryogen for plunge freezing. Microsc. Micoanal. 14, 375–379 (2008).

    Article  CAS  Google Scholar 

  34. Iancu, C. V. et al. A 'flip-flop' rotation stage for routine dual-axis electron cryotomography. J. Struct. Biol. 151, 288–297 (2005).

    Article  Google Scholar 

  35. Zheng, Q. S., Braunfeld, M. B., Sedat, J. W. & Agard, D. A. An improved strategy for automated electron microscopic tomography. J. Struct. Biol. 147, 91–101 (2004).

    Article  Google Scholar 

  36. Suloway, C. et al. Fully automated, sequential tilt-series acquisition with Leginon. J. Struct. Biol. 167, 11–18 (2009).

    Article  CAS  Google Scholar 

  37. Mastronarde, D. N. Dual-axis tomography: an approach with alignment methods that preserve resolution. J. Struct. Biol. 120, 343–352 (1997).

    Article  CAS  Google Scholar 

  38. Amat, F. et al. Markov random field based automatic image alignment for electron tomography. J. Struct. Biol. 161, 260–275 (2008).

    Article  Google Scholar 

  39. Domian, I. J., Quon, K. C. & Shapiro, L. Cell type-specific phosphorylation and proteolysis of a transcriptional regulator controls the G1-to-S transition in a bacterial cell cycle. Cell 90, 415–424 (1997).

    Article  CAS  Google Scholar 

  40. Simard, D., Hewitt, K. A., Lunn, F., Iyengar, A. & Bearne, S. L. Limited proteolysis of Escherichia coli cytidine 5′-triphosphate synthase. Identification of residues required for CTP formation and GTP-dependent activation of glutamine hydrolysis. Eur. J. Biochem. 270, 2195–2206 (2003).

    Article  CAS  Google Scholar 

Download references


We are grateful to B. Bassler, C. Murphy, E. Klein and K. Cowles for critical reading of the manuscript. We thank M. Cabeen, C. Jacobs-Wagner, J. Williamson, Alison Michaelis and other members of the Gitai lab for reagents and discussions. J.N.W. is supported by a postdoctoral fellowship from the National Institute of Allergy and Infectious Diseases (grant 1F32AI073043–01A1). A.B. and G.J.J. were supported in part by the National Institutes of Health (NIH) grant R01 AI067548, the Howard Hughes Medical Institute, and a gift to Caltech from the Gordon and Betty Moore Foundation. Z.G. is supported by funding from the U.S. Department of Energy Office of Science (Biological and Environmental Research; grant DE-FG02-05ER64136), NIH grant 1DP2OD004389-01, the Human Frontiers Science Program and the Beckman Foundation.

Author information

Authors and Affiliations



M.J.I. performed all experiments except for the ECT and fLM-ECT experiments, which were performed by A.B. J.N.W. performed the initial screen that identified CtpS as a linearly localized protein. Experiments were conceived by M.J.I., A.B., G.J.J. and Z.G., and the manuscript was written by M.J.I. and Z.G. with significant input from A.B. and G.J.J.

Corresponding author

Correspondence to Zemer Gitai.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary Information (PDF 1851 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Ingerson-Mahar, M., Briegel, A., Werner, J. et al. The metabolic enzyme CTP synthase forms cytoskeletal filaments. Nat Cell Biol 12, 739–746 (2010).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing