Skip to main content

Thank you for visiting nature.com. 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.

BAP31 and BiP are essential for dislocation of SV40 from the endoplasmic reticulum to the cytosol

Abstract

How non-enveloped viruses overcome host cell membranes is poorly understood. Here, we show that after endocytosis and transport to the endoplasmic reticulum (ER), but before crossing the ER membrane to the cytosol, incoming simian virus 40 particles are structurally remodelled leading to exposure of the amino-terminal sequence of the minor viral protein VP2. These hydrophobic sequences anchor the virus to membranes. A negatively charged residue, Glu 17, in the α-helical, membrane-embedded peptide is essential for infection, most likely by introducing an ‘irregularity’ recognized by the ER-associated degradation (ERAD) system for membrane proteins. Using a siRNA-mediated screen, the lumenal chaperone BiP and the ER-membrane protein BAP31 (both involved in ERAD) were identified as being essential for infection. They co-localized with the virus in discrete foci and promoted its ER-to-cytosol dislocation. Virus-like particles devoid of VP2 failed to cross the membrane. The results demonstrated that ERAD-factors assist virus transport across the ER membrane.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: SV40 undergoes a structural change in the ER.
Figure 2: The N terminus of VP2 folds into an α-helix, integrates into the ER membrane and contains an essential residue, Glu 17.
Figure 3: SV40 infection depends on BAP31, BAP29 and RMA1.
Figure 4: Transport of SV40 to the ER is not affected following knockdown of BAP31.
Figure 5: BAP31, BAP29 and SV40 accumulate in discrete spots in the ER.
Figure 6: BAP31 interacts with the N-terminal peptide of VP2.
Figure 7: BiP is a critical factor in SV40 infection.
Figure 8: BAP31 and BiP are essential for the transport of SV40 to the cytosol.

References

  1. 1

    Jiang, M., Abend, J. R., Johnson, S. F. & Imperiale, M. J. The role of polyomaviruses in human disease. Virology 384, 266–273 (2009).

    CAS  Article  Google Scholar 

  2. 2

    Major, E. O. Progressive multifocal leukoencephalopathy in patients on immunomodulatory therapies. Annu. Rev. Med. 61, 35–47 (2010).

    CAS  Article  Google Scholar 

  3. 3

    Eash, S., Manley, K., Gasparovic, M., Querbes, W. & Atwood, W. J. The human polyomaviruses. Cell Mol. Life Sci. 63, 865–876 (2006).

    CAS  Article  Google Scholar 

  4. 4

    Liddington, R. C. et al. Structure of simian virus 40 at 3.8-A resolution. Nature 354, 278–284 (1991).

    CAS  Article  Google Scholar 

  5. 5

    Stehle, T., Gamblin, S. J., Yan, Y. & Harrison, S. C. The structure of simian virus 40 refined at 3.1 A resolution. Structure 4, 165–182 (1996).

    CAS  Article  Google Scholar 

  6. 6

    Schelhaas, M. et al. Simian Virus 40 depends on ER protein folding and quality control factors for entry into host cells. Cell 131, 516–529 (2007).

    CAS  Article  Google Scholar 

  7. 7

    Daniels, R., Rusan, N. M., Wadsworth, P. & Hebert, D. N. SV40 VP2 and VP3 insertion into ER membranes is controlled by the capsid protein VP1: implications for DNA translocation out of the ER. Mol. Cell 24, 955–66 (2006).

    CAS  Article  Google Scholar 

  8. 8

    Streuli, C. H. & Griffin, B. E. Myristic acid is coupled to a structural protein of polyoma virus and SV40. Nature 326, 619–622 (1987).

    CAS  Article  Google Scholar 

  9. 9

    Varshavsky, A. J. et al. Compact form of SV40 viral minichromosome is resistant to nuclease: possible implications for chromatin structure. Nucleic Acids Res. 4, 3303–3325 (1977).

    CAS  Article  Google Scholar 

  10. 10

    Tsai, B. et al. Gangliosides are receptors for murine polyoma virus and SV40. EMBO J. 22, 4346–4355 (2003).

    CAS  Article  Google Scholar 

  11. 11

    Kartenbeck, J., Stukenbrok, H. & Helenius, A. Endocytosis of simian virus 40 into the endoplasmic reticulum. J. Cell Biol. 109, 2721–2729 (1989).

    CAS  Article  Google Scholar 

  12. 12

    Ewers, H. et al. GM1 structure determines SV40-induced membrane invagination and infection. Nat. Cell Biol. 12, 11–18 (2010).

    CAS  Article  Google Scholar 

  13. 13

    Neu, U., Stehle, T. & Atwood, W. J. The polyomaviridae: contributions of virus structure to our understanding of virus receptors and infectious entry. Virology 384, 389–399 (2009).

    CAS  Article  Google Scholar 

  14. 14

    Engel, S. et al. Role of endosomes in simian virus 40 entry and infection. J. Virol. 85, 4198–4211 (2011).

    CAS  Article  Google Scholar 

  15. 15

    Nakanishi, A., Clever, J., Yamada, M., Li, P. P. & Kasamatsu, H. Association with capsid proteins promotes nuclear targeting of simian virus 40 DNA. Proc. Natl Acad. Sci. USA 93, 96–100 (1996).

    CAS  Article  Google Scholar 

  16. 16

    Nakanishi, A., Shum, D., Morioka, H., Otsuka, E. & Kasamatsu, H. Interaction of the Vp3 nuclear localization signal with the importin α2/β heterodimer directs nuclear entry of infecting simian virus 40. J. Virol. 76, 9368–9377 (2002).

    CAS  Article  Google Scholar 

  17. 17

    Tsai, B. & Qian, M. Cellular entry of polyomaviruses. Curr. Top. Microbiol. Immunol. 343, 177–194 (2010).

    CAS  PubMed  Google Scholar 

  18. 18

    Ploegh, H. L. A lipid-based model for the creation of an escape hatch from the endoplasmic reticulum. Nature 448, 435–438 (2007).

    CAS  Article  Google Scholar 

  19. 19

    Vembar, S. S. & Brodsky, J. L. One step at a time: endoplasmic reticulum-associated degradation. Nat. Rev. Mol. Cell Biol. 9, 944–957 (2008).

    CAS  Article  Google Scholar 

  20. 20

    Hegde, R. S. & Ploegh, H. L. Quality and quantity control at the endoplasmic reticulum. Curr. Opin. Cell Biol. 22, 437–446 (2010).

    CAS  Article  Google Scholar 

  21. 21

    Hirsch, C., Gauss, R., Horn, S. C., Neuber, O. & Sommer, T. The ubiquitylation machinery of the endoplasmic reticulum. Nature 458, 453–460 (2009).

    CAS  Article  Google Scholar 

  22. 22

    Carvalho, P., Goder, V. & Rapoport, T. A. Distinct ubiquitin-ligase complexes define convergent pathways for the degradation of ER proteins. Cell 126, 361–373 (2006).

    CAS  Article  Google Scholar 

  23. 23

    Xie, W. & Ng, D. T. ERAD substrate recognition in budding yeast. Semin. CellDev. Biol. 21, 533–539 (2010).

    CAS  Article  Google Scholar 

  24. 24

    Tsai, B., Rodighiero, C., Lencer, W. I. & Rapoport, T. A. Protein disulfide isomerase acts as a redox-dependent chaperone to unfold cholera toxin. Cell 104, 937–948 (2001).

    CAS  Article  Google Scholar 

  25. 25

    Magnuson, B. et al. ERp29 triggers a conformational change in polyomavirus to stimulate membrane binding. Mol. Cell 20, 289–300 (2005).

    CAS  Article  Google Scholar 

  26. 26

    Chen, X. S., Stehle, T. & Harrison, S. C. Interaction of polyomavirus internal protein VP2 with the major capsid protein VP1 and implications for participation of VP2 in viral entry. EMBO J. 17, 3233–3240 (1998).

    CAS  Article  Google Scholar 

  27. 27

    Bonifacino, J. S., Cosson, P., Shah, N. & Klausner, R. D. Role of potentially charged transmembrane residues in targeting proteins for retention and degradation within the endoplasmic reticulum. EMBO J. 10, 2783–2793 (1991).

    CAS  Article  Google Scholar 

  28. 28

    Ray-Sinha, A., Cross, B. C., Mironov, A., Wiertz, E. & High, S. Endoplasmic reticulum-associated degradation of a degron-containing polytopic membrane protein. Mol. Membr. Biol. 26, 448–464 (2009).

    Article  Google Scholar 

  29. 29

    Keppler, A. et al. A general method for the covalent labeling of fusion proteins with small molecules in vivo. Nat. Biotechnol. 21, 86–89 (2003).

    CAS  Article  Google Scholar 

  30. 30

    Shibata, Y. et al. Mechanisms determining the morphology of the peripheral ER. Cell 143, 774–788 (2010).

    CAS  Article  Google Scholar 

  31. 31

    Schamel, W. W. et al. A high-molecular-weight complex of membrane proteins BAP29/BAP31 is involved in the retention of membrane-bound IgD in the endoplasmic reticulum. Proc. Natl Acad. Sci. USA 100, 9861–9866 (2003).

    CAS  Article  Google Scholar 

  32. 32

    Wang, B. et al. BAP31 interacts with Sec61 translocons and promotes retrotranslocation of CFTRΔF508 via the derlin-1 complex. Cell 133, 1080–1092 (2008).

    CAS  Article  Google Scholar 

  33. 33

    Wakana, Y. et al. Bap31 is an itinerant protein that moves between the peripheral endoplasmic reticulum (ER) and a juxtanuclear compartment related to ER-associated degradation. Mol. Biol. Cell 19, 1825–1836 (2008).

    CAS  Article  Google Scholar 

  34. 34

    Lilley, B. N. & Ploegh, H. L. A membrane protein required for dislocation of misfolded proteins from the ER. Nature 429, 834–840 (2004).

    CAS  Article  Google Scholar 

  35. 35

    Ye, Y., Shibata, Y., Yun, C., Ron, D. & Rapoport, T. A. A membrane protein complex mediates retro-translocation from the ER lumen into the cytosol. Nature 429, 841–847 (2004).

    CAS  Article  Google Scholar 

  36. 36

    Younger, J. M. et al. Sequential quality-control checkpoints triage misfolded cystic fibrosis transmembrane conductance regulator. Cell 126, 571–582 (2006).

    CAS  Article  Google Scholar 

  37. 37

    Kikkert, M. et al. Human HRD1 is an E3 ubiquitin ligase involved in degradation of proteins from the endoplasmic reticulum. J. Biol. Chem. 279, 3525–3534 (2004).

    CAS  Article  Google Scholar 

  38. 38

    Bernasconi, R., Galli, C., Calanca, V., Nakajima, T. & Molinari, M. Stringent requirement for HRD1, SEL1L, and OS-9/XTP3-B for disposal of ERAD-LS substrates. J. Cell Biol. 188, 223–235 (2010).

    CAS  Article  Google Scholar 

  39. 39

    Okuda-Shimizu, Y. & Hendershot, L. M. Characterization of an ERAD pathway for nonglycosylated BiP substrates, which require Herp. Mol. Cell 28, 544–554 (2007).

    CAS  Article  Google Scholar 

  40. 40

    Oda, Y., Hosokawa, N., Wada, I. & Nagata, K. EDEM as an acceptor of terminally misfolded glycoproteins released from calnexin. Science 299, 1394–1397 (2003).

    CAS  Article  Google Scholar 

  41. 41

    Molinari, M., Calanca, V., Galli, C., Lucca, P. & Paganetti, P. Role of EDEM in the release of misfolded glycoproteins from the calnexin cycle. Science 299, 1397–1400 (2003).

    CAS  Article  Google Scholar 

  42. 42

    Ye, Y., Meyer, H. H. & Rapoport, T. A. The AAA ATPase Cdc48/p97 and its partners transport proteins from the ER into the cytosol. Nature 414, 652–656 (2001).

    CAS  Article  Google Scholar 

  43. 43

    Ye, Y., Meyer, H. H. & Rapoport, T. A. Function of the p97-Ufd1-Npl4complex in retrotranslocation from the ER to the cytosol: dual recognition of nonubiquitinated polypeptide segments and polyubiquitin chains. J. Cell Biol. 162, 71–84 (2003).

    CAS  Article  Google Scholar 

  44. 44

    Carlson, E. J., Pitonzo, D & Skach, W. R. p97 functions as an auxiliary factor to facilitate TM domain extraction during CFTR ER-associated degradation. EMBO J. 25, 4557–4566 (2006).

    CAS  Article  Google Scholar 

  45. 45

    Nowis, D., McConnell, E. & Wojcik, C. Destabilization of the VCP-Ufd1-Npl4 complex is associated with decreased levels of ERAD substrates. Exp. Cell Res. 312, 2921–2932 (2006).

    CAS  Article  Google Scholar 

  46. 46

    Ng, F. W. et al. p28 Bap31, a Bcl-2/Bcl-XL- and procaspase-8-associated protein in the endoplasmic reticulum. J. Cell Biol. 139, 327–338 (1997).

    CAS  Article  Google Scholar 

  47. 47

    Breckenridge, D. G., Nguyen, M., Kuppig, S., Reth, M. & Shore, G. C. The procaspase-8 isoform, procaspase-8L, recruited to the BAP31 complex at the endoplasmic reticulum. Proc. Natl Acad. Sci. USA 99, 4331–4336 (2002).

    CAS  Article  Google Scholar 

  48. 48

    Romisch, K. Endoplasmic reticulum-associated degradation. Annu. Rev. CellDev. Biol. 21, 435–456 (2005).

    CAS  Article  Google Scholar 

  49. 49

    Nishikawa, S. I., Fewell, S. W., Kato, Y., Brodsky, J. L. & Endo, T. Molecular chaperones in the yeast endoplasmic reticulum maintain the solubility of proteins for retrotranslocation and degradation. J. Cell Biol. 153, 1061–1070 (2001).

    CAS  Article  Google Scholar 

  50. 50

    Blond-Elguindi, S. et al. Affinity panning of a library of peptides displayed on bacteriophages reveals the binding specificity of BiP. Cell 75, 717–728 (1993).

    CAS  Article  Google Scholar 

  51. 51

    Bernardi, K. M., Forster, M. L., Lencer, W. I. & Tsai, B. Derlin-1 facilitates the retro-translocation of cholera toxin. Mol. Biol. Cell 19, 877–884 (2008).

    CAS  Article  Google Scholar 

  52. 52

    Richards, A. A., Stang, E., Pepperkok, R. & Parton, R. G. Inhibitors of COP-mediated transport and cholera toxin action inhibit simian virus 40 infection. Mol. Biol. Cell 13, 1750–1764 (2002).

    CAS  Article  Google Scholar 

  53. 53

    Kamhi-Nesher, S. et al. A novel quality control compartment derived from the endoplasmic reticulum. Mol. Biol. Cell 12, 1711–1723 (2001).

    CAS  Article  Google Scholar 

  54. 54

    Norkin, L. C., Anderson, H. A., Wolfrom, S. A. & Oppenheim, A. Caveolar endocytosis of simian virus 40 is followed by brefeldin A-sensitive transport to the endoplasmic reticulum, where the virus disassembles. J. Virol. 76, 5156–5166 (2002).

    CAS  Article  Google Scholar 

  55. 55

    Engelman, D. M. Electrostatic fasteners hold the T cell receptor-CD3 complex together. Mol. Cell 11, 5–6 (2003).

    CAS  Article  Google Scholar 

  56. 56

    Gilbert, J., Ou, W., Silver, J & Benjamin, T. Downregulation of protein disulfide isomerase inhibits infection by the mouse polyomavirus. J. Virol. 80, 10868–10870 (2006).

    CAS  Article  Google Scholar 

  57. 57

    Lilley, B. N., Gilbert, J. M., Ploegh, H.L. & Benjamin, T. L. Murine polyomavirus requires the endoplasmic reticulum protein Derlin-2 to initiate infection. J. Virol. 80, 8739–8744 (2006).

    CAS  Article  Google Scholar 

  58. 58

    Pelkmans, L., Kartenbeck, J. & Helenius, A. Caveolar endocytosis of simian virus 40 reveals a new two-step vesicular-transport pathway to the ER. Nat. Cell Biol. 3, 473–483 (2001).

    CAS  Article  Google Scholar 

  59. 59

    Ishii, N., Nakanishi, A., Yamada, M., Macalalad, M. H. & Kasamatsu, H. Functional complementation of nuclear targeting-defective mutants of simian virus 40 structural proteins. J. Virol. 68, 8209–8216 (1994).

    CAS  PubMed  PubMed Central  Google Scholar 

  60. 60

    Walter, P. & Blobel, G. Preparation of microsomal membranes for cotranslational protein translocation. Methods Enzymol. 96, 84–93 (1983).

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We thank H. Ploegh, G. Shore, E. Wiertz, H. Meyer, H. Kasamatsu, A. Oppenheim, L. Hendershot and K. Johnsson for sharing reagents. We are grateful to A. Smith, P. Deprez, J. Kartenbeck and R. Mancini for preparing mPy and microsomes and for help with electron microscopy. We thank D. Roderer for help with circular dichroism spectroscopy measurements. We thank members of the Helenius group for discussions, especially M. Schelhaas. This work was supported by the National Science Foundation of Switzerland, the LipidX programme of SystemsX, the European research council (ERC) and ETH Zurich.

Author information

Affiliations

Authors

Contributions

R.G. designed and carried out experiments and analysed the data. D.A., S.F., F.H. and S.L. carried out experiments. T.H. wrote the MATLAB program. R.G. and A.H. wrote the manuscript. A.H. supervised the work.

Corresponding author

Correspondence to Ari Helenius.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary Information (PDF 1783 kb)

Supplementary Table 1

Supplementary Information (XLS 100 kb)

Supplementary Table 2

Supplementary Information (XLSX 38 kb)

Supplementary Table 3

Supplementary Information (XLS 30 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Geiger, R., Andritschke, D., Friebe, S. et al. BAP31 and BiP are essential for dislocation of SV40 from the endoplasmic reticulum to the cytosol. Nat Cell Biol 13, 1305–1314 (2011). https://doi.org/10.1038/ncb2339

Download citation

Further reading

Search

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