Abstract
The post-genome packaging mechanisms that govern the assembly of an infectious virion are poorly understood in bacteriophages and other viruses. Here, our near-atomic resolution cryo-EM structural analyses uncovered an assembly- and conformation-driven genome positioning mechanism in the tailed bacteriophage T4. We show that following headful packaging, which generates a pressurized head, a global conformational change occurs in the portal structure, probably triggering packaging termination and ejection of the packaging motor. Our high-resolution structures of the neck of the virion further show that the neck undergoes conformational changes upon docking of a pre-assembled tail onto the sealed neck, which then opens a genome-gate. Driven by the pressure of the packaged DNA, the genome travels through open neck channels, binds and compresses the resident tape-measure protein, and halts at the bottom of the second topmost disk of the tail tube. Pressure-suspended within the virion’s innermost tunnel and secured by a baseplate plug, the genome remains poised to flow through the tunnel into a host cell upon receiving the host receptor recognition signal.
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Data availability
The C1 and C6 symmetric cryo-EM reconstructions focused on the phage neck region have been deposited in the EM Data Bank with the accession codes EMD-48458 and EMD-48462, respectively. The C3 and C6 symmetric reconstructions of the middle part of the tail have been deposited with the accession codes EMD-48460 and EMD-48463, respectively; and the C3 and C6 symmetric reconstructions of the distal part of the tail have been deposited with the accession codes EMD-48459 and EMD-48464, respectively. The composite map of the whole portal-neck-tail complex has been deposited with the accession code EMD-48324. The asymmetric atomic structure of the neck has been deposited in the Protein Data Bank (PDB) with the accession code 9MOF. The three-fold-symmetric structures of the middle and distal parts of the tail have been deposited with the PDB accession codes 9MOH and 9MOG, respectively. The composite structure of the whole portal-neck-tail complex has been deposited with the PDB accession code 9MKB.
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Acknowledgements
This work was supported by NIAID, NIH grant 1R01AI175340 to V.B.R. and A.F. and in part by NIDA, NIH Avant Garde Award 1DP1DA060580 and National Science Foundation grant MCB-0923873 to V.B.R. Research in Q.F.’s laboratory was supported by the National Natural Science Foundation of China (32371285).
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V.B.R., M.G.R., and A.F. conceived the project. A.F., J.Z., T.K., F.V., C-A.A., Z.W., B.K., and V.B.R. performed research. A.F., J.Z., C-A.A., M.G.R., Z.C., L.S., Q.F., R.J.K. and V.B.R. analyzed data. V.B.R., R.J.K., and M.G.R. supervised the project. A.F. and V.B.R. prepared the manuscript, with additional edits from all authors. V.B.R. provided overall direction and coordination for this and the accompanying genome retention project.
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Fokine, A., Zhu, J., Klose, T. et al. In situ structures of the portal-neck-tail complex of bacteriophage T4 inform a viral genome positioning mechanism. Nat Commun (2026). https://doi.org/10.1038/s41467-026-69106-8
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DOI: https://doi.org/10.1038/s41467-026-69106-8
