PDB 4V60: structure of a protein vault (MVP only)
Structure and organization:
Wild-type vaults consist of multiple copies of major vault protein (MVP), VPARP, and TEP1 proteins as well as small untranslated RNAs called vRNAs (Pupols, 2011).
Vaults are ~13 MDa in mass if VPARP, TEP1, and vRNAs are included along with the 78 MVPs (Galbiati et al., 2018). Each of the 78 MVP copies is ~97 kDa (Champion et al., 2009), so the hollow vault mass is ~7.7 MDa.
VPARP catalyzes poly-ADP ribosylation. It has been found to ribosylate itself and MVP. Its function is unknown. It contains the INT domain, which binds to the interior waist region of the vault (Pupols, 2011).
TEP1 (telomerase-associated protein 1) is found both associated with nuclear telomerase complexes and with cytosolic vaults. Its function is unknown (Pupols, 2011).
vRNAs are untranslated RNA polymerase III transcripts ranging from 80-150 nucleotides in length. Their function is unknown (Pupols, 2011).
Physiology and dynamics:
Vaults are found inside every cell in the human body at copy numbers of ~104 vaults per cell in most cells but ~105 in certain cell types (e.g. some immune cells) (Travis, 2024). They are especially abundant in tissues exposed to external stressors such as bronchus, renal proximal tubules, digestive tract, macrophages, and dendritic cells (Pupols, 2011). In embryonic tissues, vaults sometimes can occur at an impressive ~107 copies per cell (Suprenant, 2002).
Vaults do not self-assemble from MVP on its own. They can only be made co-translationally on eukaryotic polyribosomes (Mrazek et al., 2014). Two copies of MVP, oriented in opposite directions, are first translated by two ribosomes. The N-terminal regions of these MVPs dimerize. As more ribosome pairs arrive in line, more MVP dimers are made. Lateral interactions between the dimers begin assembling the wall of the vault. In total, 39 copies of MVP dimer are translated on the polyribosome, leading to the formation of the final barrel-shaped vault structure.
Vaults have consistently been found as contaminants in purified extracellular vesicle (EV) preparations. There is evidence that vaults associate with the outside of EVs and are not protected beneath vesicular membranes (Liu et al., 2023). However, vaults have also been found to be released from cells in an EV-independent fashion wherein they are not bound to the outside of the EVs (Jeppesen et al., 2019). As such, they might be co-released alongside EVs.
Vaults frequently exchange halves when in solution, indicating their dynamic structural nature (Yang et al., 2010). Indeed, vaults have been proposed to experience a structural “breathing” motion.
Vaults disassemble at low pH, through mechanisms of half vault separation (Goldsmith et al., 2007) and/or weakening of the lateral associations between MVP copies (Llauró et al., 2016).
Vaults are cytosolic particles, but small amounts of them associate with the nuclear membrane at nuclear pore complexes and in some cell types (e.g. U373 glioblastoma cell line) about 5% of MVP is localized to the nucleus (Slesina et al., 2005).
As MVP is a self-protein, it is usually invisible to the immune system (Champion et al., 2009). Indeed, repeated intranasal administration of vaults carrying non-immunogenic proteins like mCherry-INT does not induce anti-vault antibodies even when MVP is fused to Z peptide (an Fc-binding peptide often used to conjugate antibodies for vault targeting to specific cell types). That said, the immune system’s tolerance to vaults can be broken if vaults carrying highly immunogenic proteins like chlamydia major outer membrane protein with INT (MOMP-INT). Repeated administration of vaults carrying MOMP-INT has been shown to induce anti-MVP antibodies.
Hints at function:
MVP knockout mice are viable but have lower survival rates when challenged with Pseudomonas aeruginosa (Frascotti et al., 2021).
Vault overexpression is found in multidrug resistant cancers, but so far this seems more of a correlation than a causation. Experimentally, overexpression of vaults alone does not produce the multidrug resistant phenotype (Frascotti et al., 2021).
In neurons, vaults localize at neurite tips and along axonal and dendritic microtubule networks. Vaults can co-precipitate with cytoplasmic RNAs that are known to be translated in response to synaptic activity (Frascotti et al., 2021).
Vaults are highly conserved (Daly et al., 2013; Slinning et al., 2024). They occur in mammals, amphibians, birds, fish, sea urchins, slime molds, and more. That said, insects, plants, and fungi do not have vaults.
References:
Champion, C. I., Kickhoefer, V. A., Liu, G., Moniz, R. J., Freed, A. S., Bergmann, L. L., Vaccari, D., Raval-Fernandes, S., Chan, A. M., Rome, L. H., & Kelly, K. A. (2009). A Vault Nanoparticle Vaccine Induces Protective Mucosal Immunity. PLOS ONE, 4(4), e5409. https://doi.org/10.1371/journal.pone.0005409
Daly, T. K., Sutherland-Smith, A. J., & Penny, D. (2013). In Silico Resurrection of the Major Vault Protein Suggests It Is Ancestral in Modern Eukaryotes. Genome Biology and Evolution, 5(8), 1567–1583. https://doi.org/10.1093/gbe/evt113
Frascotti, G., Galbiati, E., Mazzucchelli, M., Pozzi, M., Salvioni, L., Vertemara, J., & Tortora, P. (2021). The Vault Nanoparticle: A Gigantic Ribonucleoprotein Assembly Involved in Diverse Physiological and Pathological Phenomena and an Ideal Nanovector for Drug Delivery and Therapy. In Cancers (Vol. 13, Issue 4). https://doi.org/10.3390/cancers13040707
Galbiati, E., Avvakumova, S., La Rocca, A., Pozzi, M., Messali, S., Magnaghi, P., Colombo, M., Prosperi, D., & Tortora, P. (2018). A fast and straightforward procedure for vault nanoparticle purification and the characterization of its endocytic uptake. Biochimica et Biophysica Acta (BBA) – General Subjects, 1862(10), 2254–2260. https://doi.org/https://doi.org/10.1016/j.bbagen.2018.07.018
Goldsmith, L. E., Yu, M., Rome, L. H., & Monbouquette, H. G. (2007). Vault Nanocapsule Dissociation into Halves Triggered at Low pH. Biochemistry, 46(10), 2865–2875. https://doi.org/10.1021/bi0606243
Jeppesen, D. K., Fenix, A. M., Franklin, J. L., Higginbotham, J. N., Zhang, Q., Zimmerman, L. J., Liebler, D. C., Ping, J., Liu, Q., Evans, R., Fissell, W. H., Patton, J. G., Rome, L. H., Burnette, D. T., & Coffey, R. J. (2019). Reassessment of Exosome Composition. Cell, 177(2), 428-445.e18. https://doi.org/10.1016/j.cell.2019.02.029
Liu, X., Nizamudeen, Z., Hill, C. J., Parmenter, C., Arkill, K. P., Lambert, D. W., & Hunt, S. (2023). Vault particles are common contaminants of extracellular vesicle preparations. BioRxiv, 2023.11.09.566362. https://doi.org/10.1101/2023.11.09.566362
Llauró, A., Guerra, P., Kant, R., Bothner, B., Verdaguer, N., & de Pablo, P. J. (2016). Decrease in pH destabilizes individual vault nanocages by weakening the inter-protein lateral interaction. Scientific Reports, 6(1), 34143. https://doi.org/10.1038/srep34143
Mrazek, J., Toso, D., Ryazantsev, S., Zhang, X., Zhou, Z. H., Fernandez, B. C., Kickhoefer, V. A., & Rome, L. H. (2014). Polyribosomes Are Molecular 3D Nanoprinters That Orchestrate the Assembly of Vault Particles. ACS Nano, 8(11), 11552–11559. https://doi.org/10.1021/nn504778h
Pupols, M. D. (2011). Packaging RNA into Vault Nanoparticles to Develop a Novel Delivery System for RNA Therapeutics. In ProQuest Dissertations and Theses. University of California, Los Angeles PP – United States — California.
Slesina, M., Inman, E. M., Rome, L. H., & Volknandt, W. (2005). Nuclear localization of the major vault protein in U373 cells. Cell and Tissue Research, 321(1), 97–104. https://doi.org/10.1007/s00441-005-1086-8
Slinning, M. S., Nthiga, T. M., Eichner, C., Khadija, S., Rome, L. H., Nilsen, F., & Dondrup, M. (2024). Major vault protein is part of an extracellular cement material in the Atlantic salmon louse (Lepeophtheirus salmonis). Scientific Reports, 14(1), 15240. https://doi.org/10.1038/s41598-024-65683-0
Suprenant, K. A. (2002). Vault Ribonucleoprotein Particles: Sarcophagi, Gondolas, or Safety Deposit Boxes? Biochemistry, 41(49), 14447–14454. https://doi.org/10.1021/bi026747e
Travis, J. (2024). The vault guy. Science (New York, NY), 384(6700), 1058–1062.
Yang, J., Kickhoefer, V. A., Ng, B. C., Gopal, A., Bentolila, L. A., John, S., Tolbert, S. H., & Rome, L. H. (2010). Vaults Are Dynamically Unconstrained Cytoplasmic Nanoparticles Capable of Half Vault Exchange. ACS Nano, 4(12), 7229–7240. https://doi.org/10.1021/nn102051r
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