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Spatial distribution and structural arrangement of a murine cytomegalovirus glycoprotein detected by SPDM localization microscopy

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Abstract

Novel approaches of localization microscopy have opened new insights into the molecular nano-cosmos of cells. We applied a special embodiment called spectral position determination microscopy (SPDM) that has the advantage to run with standard fluorescent dyes or proteins under standard preparation conditions. Pointillist images with a resolution in the order of 10 nm can be obtained by SPDM. Therefore, vector pEYFP-m164, encoding the murine cytomegalovirus glycoprotein gp36.5/m164 fused to enhanced yellow fluorescent protein, was transiently transfected into COS-7 cells. This protein shows exceptional intracellular trafficking dynamics, moving within the endoplasmic reticulum (ER) and outer nuclear membrane. The molecular positions of gp36.5/m164 were visualized and determined by SPDM imaging. From the position point patterns of the protein molecules, their arrangements were quantified by next neighbour distance analyses. Three different structural arrangements were discriminated: (a) a linear distribution along the membrane, (b) a highly structured distribution in the ER, and (c) a homogenous distribution in the cellular cytoplasm. The results indicate that the analysis of next neighbour distances on the nano-scale allows the identification and discrimination of different structural arrangements of molecules within their natural cellular environment.

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Acknowledgments

The authors gratefully acknowledge the financial support of the Deutsche Forschungsgemeinschaft (German Research Council) to C.C., and of the Bundesministerium für Bildung und Forschung (Federal Ministry for Education and Research) to M.H., R.H. was supported by the Deutsche Forschungsgemeinschaft, Collaborative Research Centre (SFB) 490 (TP-E3) and the intramural funding in program IFF-I of the University Medical Center, Mainz, Germany. N.A.L. received intramural funding in the young investigator program MAIFOR of the University Medical Center, Mainz, Germany. The authors thank Udo Birk, IMB, Mainz, for technical contributions in the development of the SPDM instrumentation.

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Author notes

  1. Rainer Kaufmann

    Present address: Division of Structural Biology, Wellcome Trust Centre for Human Genetics and Department of Biochemistry, University of Oxford, Oxford, UK

  2. Manuel Gunkel

    Present address: Bioquant, University of Heidelberg, Heidelberg, Germany

  3. Daniel Paech

    Present address: Division of Neuroradiology, University Medical Center Heidelberg, Heidelberg, Germany

Authors and Affiliations

  1. Kirchhoff-Institute for Physics, Im Neuenheimer Feld 227, 69120, Heidelberg, Germany

    Patrick Müller, Rainer Kaufmann, Manuel Gunkel, Daniel Paech, Georg Hildenbrand, Christoph Cremer & Michael Hausmann

  2. Institute for Virology, University Medical Center Mainz, Mainz, Germany

    Niels A. Lemmermann & Rafaela Holtappels

  3. Department of Radiation Oncology, University Medical Center Mannheim, Mannheim, Germany

    Georg Hildenbrand

  4. Institute for Pharmacy and Molecular Biotechnology, University Heidelberg, Heidelberg, Germany

    Christoph Cremer

  5. Institute of Molecular Biology (IMB), Mainz, Germany

    Christoph Cremer

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  1. Patrick Müller
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Correspondence to Michael Hausmann.

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Müller, P., Lemmermann, N.A., Kaufmann, R. et al. Spatial distribution and structural arrangement of a murine cytomegalovirus glycoprotein detected by SPDM localization microscopy. Histochem Cell Biol 142, 61–67 (2014). https://doi.org/10.1007/s00418-014-1185-2

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