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.

Two DNA nanomachines map pH changes along intersecting endocytic pathways inside the same cell


DNA is a versatile scaffold for molecular sensing in living cells, and various cellular applications of DNA nanodevices have been demonstrated. However, the simultaneous use of different DNA nanodevices within the same living cell remains a challenge. Here, we show that two distinct DNA nanomachines can be used simultaneously to map pH gradients along two different but intersecting cellular entry pathways. The two nanomachines, which are molecularly programmed to enter cells via different pathways, can map pH changes within well-defined subcellular environments along both pathways inside the same cell. We applied these nanomachines to probe the pH of early endosomes and the trans-Golgi network, in real time. When delivered either sequentially or simultaneously, both nanomachines localized into and independently captured the pH of the organelles for which they were designed. The successful functioning of DNA nanodevices within living systems has important implications for sensing and therapies in a diverse range of contexts.

Figure 1: Programming of DNA nanodevices.
Figure 2: Programmed trafficking of DNA nanomachines along two distinct endocytic pathways.
Figure 3: In cellulo performance of DNA nanomachines.
Figure 4: SimpHony of transferrin receptor and furin endocytic pathways.
Figure 5: SimpHony of organelles with altered morphology.


  1. 1

    Seeman, N. C. DNA in a material world. Nature 421, 427–431 (2003).

    Article  Google Scholar 

  2. 2

    Bath, J. & Turberfield, A. J. DNA nanomachines. Nature Nanotech. 2, 275–284 (2007).

    CAS  Article  Google Scholar 

  3. 3

    Krishnan, Y. & Bathe, M. Designer nucleic acids to probe and program the cell. Trends Cell Biol. 22, 624–633 (2012).

    CAS  Article  Google Scholar 

  4. 4

    McMahon, D. Chemical messengers in development: a hypothesis. Science 185, 1012–1021 (1974).

    CAS  Article  Google Scholar 

  5. 5

    Modi, S. et. al. A DNA nanomachine that maps spatial and temporal pH changes inside living cells. Nature Nanotech. 4, 325–330 (2009).

    CAS  Article  Google Scholar 

  6. 6

    Surana, S., Bhat, J. M., Koushika, S. P. & Krishnan, Y. An autonomous DNA nanomachine maps spatiotemporal pH changes in a multicellular living organism. Nature Commun. 2, 340 (2011).

    Article  Google Scholar 

  7. 7

    Douglas, S. M., Bachelet, I. & Church, G. M. A logic-gated nanorobot for targeted transport of molecular payloads. Science 335, 831–834 (2012).

    CAS  Article  Google Scholar 

  8. 8

    Lee, H. et al. Molecularly self-assembled nucleic acid nanoparticles for targeted in vivo siRNA delivery. Nature Nanotech. 7, 389–393 (2012).

    CAS  Article  Google Scholar 

  9. 9

    Bhatia, D., Surana, S., Chakraborty, S., Koushika, S. P. & Krishnan, Y. A synthetic icosahedral DNA-based host–cargo complex for functional in vivo imaging. Nature Commun. 2, 339 (2011).

    Article  Google Scholar 

  10. 10

    Mallet, W. G. & Maxfield, F. R. Chimeric forms of furin and TGN38 are transported with the plasma membrane in the trans-Golgi network via distinct endosomal pathways. J. Cell Biol. 146, 345–359 (1999).

    CAS  Article  Google Scholar 

  11. 11

    Chia, P. Z. C., Gasnereau, I., Lieu, Z. Z. & Gleeson, P. A. Rab9-dependent retrograde transport and endosomal sorting of the endopeptidase furin. J. Cell Sci. 124, 2401–2413 (2011).

    CAS  Article  Google Scholar 

  12. 12

    Presley, J. F. et al. The End2 mutation in CHO cells slows the exit of transferrin receptors from the recycling compartment but bulk membrane recycling is unaffected. J. Cell Biol. 122, 1231–1241 (1993).

    CAS  Article  Google Scholar 

  13. 13

    McCafferty, J., Griffiths, A. D., Winter, G. & Chiswell, D. J. Phage antibodies: filamentous phage displaying antibody variable domains. Nature 348, 552–554 (1990).

    CAS  Article  Google Scholar 

  14. 14

    Geisow, M. J., D'Arcy Hart, P. & Young, M. R. Temporal changes of lysosome and phagosome pH during phagolysosome formation in macrophages: studies by fluorescence spectroscopy. J. Cell Biol. 89, 645–652 (1981).

    CAS  Article  Google Scholar 

  15. 15

    Llopis, J., McCaffery, J. M., Miyawaki, A., Farquhar, M. G. & Tsien, R. Y. Measurement of cytosolic, mitochondrial, and golgi pH in single living cells with green fluorescent proteins. Proc. Natl Acad. Sci. USA 95, 6803–6808 (1998).

    CAS  Article  Google Scholar 

  16. 16

    Kim, J. H. et al. Noninvasive measurement of the pH of the endoplasmic reticulum at rest and during calcium release. Proc. Natl Acad. Sci. USA 95, 2997–3002 (1998).

    CAS  Article  Google Scholar 

  17. 17

    Molloy, S. S., Thomas, L., VanSlyke, J. K., Stenberg, P. E. & Thomas, G. Intracellular trafficking and activation of the furin proprotein convertase: localization to the TGN and recycling from the cell surface. EMBO J. 13, 18–33 (1994).

    CAS  Article  Google Scholar 

  18. 18

    Molloy, S. S., Anderson, E. D., Jean, F. & Thomas, G. Bi-cycling the furin pathway: from TGN localization to pathogen activation and embryogenesis. Trends Cell Biol. 9, 28–35 (1999).

    CAS  Article  Google Scholar 

  19. 19

    Presley, J. F., Mayor, S., McGraw, T. E., Dunn, K. W. & Maxfield, F. R. Bafilomycin A1 treatment retards transferrin receptor recycling more than bulk membrane recycling. J. Biol. Chem. 272, 13929–13936 (1997).

    CAS  Article  Google Scholar 

  20. 20

    Yamashiro, D. J. & Maxfield, F. R. Acidification of morphologically distinct endosomes in mutant and wild-type Chinese hamster ovary cells. J. Cell Biol. 105, 2723–2733 (1987).

    CAS  Article  Google Scholar 

  21. 21

    Maeda, Y., Ide, T., Koike, M., Uchiyama, Y. & Kinoshita, T. GPHR is a novel anion channel critical for acidification and functions of the Golgi apparatus. Nature Cell Biol. 10, 1135–1145 (2008).

    CAS  Article  Google Scholar 

  22. 22

    Derivery, E. et al. The Arp2/3 Activator WASH controls the fission of endosomes through a large multiprotein complex. Dev. Cell 17, 712–723 (2009).

    CAS  Article  Google Scholar 

  23. 23

    Mesaki, K., Tanabe, K., Obayashi, M., Oe, N. & Takei, K. Fission of tubular endosomes triggers endosomal acidification and movement. PLoS ONE 6, e19764 (2011).

    CAS  Article  Google Scholar 

  24. 24

    Macia, E. et al. Dynasore, a cell-permeable inhibitor of dynamin. Dev. Cell 10, 839–850 (2006).

    CAS  Article  Google Scholar 

  25. 25

    Reaves, B. & Banting, G. Perturbation of the morphology of the trans-Golgi network following Brefeldin A treatment: redistribution of a TGN-specific integral membrane protein, TGN38. J. Cell Biol. 116, 85–94 (1992).

    CAS  Article  Google Scholar 

  26. 26

    Sciaky, N. et al. Golgi tubule traffic and the effects of brefeldin A visualized in living cells. J. Cell Biol. 139, 1137–1155 (1997).

    CAS  Article  Google Scholar 

  27. 27

    Strous, G. J. et al. Brefeldin A induces a microtubule-dependent fusion of galactosyltransferase-containing vesicles with the rough endoplasmic reticulum. Biol. Cell 71, 25–31 (1991).

    CAS  Article  Google Scholar 

  28. 28

    Waguri, S. et al. Visualization of TGN to endosome trafficking through fluorescently labeled MPR and AP-1 in living cells. Mol. Biol. Cell 14, 142–155 (2003).

    CAS  Article  Google Scholar 

  29. 29

    Maeda, Y., Beznoussenko, G. V., Van Lint, J., Mironov, A. A. & Malhotra, V. Recruitment of protein kinase D to the trans-Golgi network via the first cysteine-rich domain. EMBO J. 20, 5982–5990 (2001).

    CAS  Article  Google Scholar 

Download references


The authors thank S. Mayor, D. Lilley, A. Sarin, G.V. Shivashankar and W. Shih for critical input, and the Central Imaging and Flow Facility at NCBS for imaging. The authors also thank S. Mayor for scFv libraries and the IA2.2 cell line, and J. Bonifacino and M. Marks for the Tac-furin chimera plasmids. S.M., S.S. and S.H. thank the CSIR for research fellowships. C.N. thanks NCBS for generous support of this collaboration. Y.K. thanks the Wellcome Trust–DBT India Alliance and the Innovative Young Biotechnologist Award for funding.

Author information




S.M. and Y.K. conceived and designed the experiments. C.N. contributed phage display expertise. S.M. performed the in vitro and in cellulo experiments. S.S. optimized the IFu used herein, and S.H. addressed scFv–furin stability. S.M. and Y.K. analysed the data. S.M., S.S. and Y.K. co-wrote the paper and all authors commented on the manuscript.

Corresponding author

Correspondence to Yamuna Krishnan.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary information

Supplementary information (PDF 5833 kb)

Supplementary movie S1

Supplementary movie S1 (MP4 842 kb)

Supplementary movie S2

Supplementary movie S2 (MP4 1010 kb)

Supplementary movie S3

Supplementary movie S3 (MP4 1009 kb)

Supplementary movie S4

Supplementary movie S4 (MP4 2045 kb)

Supplementary movie S5

Supplementary movie S5 (MP4 2059 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Modi, S., Nizak, C., Surana, S. et al. Two DNA nanomachines map pH changes along intersecting endocytic pathways inside the same cell. Nature Nanotech 8, 459–467 (2013).

Download citation

Further reading


Quick links

Find nanotechnology articles, nanomaterial data and patents all in one place. Visit Nano by Nature Research