Related Experiment Videos
Mode of host cell penetration by bacteriophage phi X174
Abstract:
Bacteriophage phiX174 is an icosahedral phage which attaches to host cells without the aid of a complex tail assembly. When phiX174 was mixed with cell walls isolated from the bacterial host, the virions attached to the wall fragments and the phage deoxyribonucleic acid (DNA) was released. Attachment was prevented if the cell walls were treated with chloroform. Release of phage DNA, but not viral attachment, was prevented if the cell walls were incubated with lysozyme or if the virions were inactivated with formaldehyde. Treatment of the cell walls with lysozyme released structures which were of uniform size (6.5 by 25 nm). These structures attached phiX174 at the tip of one of its 12 vertices, but the viral DNA was not released. The virions attached to these structures were oriented with their fivefold axis of symmetry normal to the long axis of the structure. No virions were attached to these structures by more than one vertex. Freeze-etch preparations of phiX174 adsorbed to intact bacteria showed that the virions were submerged to one half their diameter into the host cell wall, and the fivefold axis of symmetry was normal to the cell surface. A second cell could not be attached to the outwardly facing vertex of the adsorbed phage and thus the phage could not cross-link two cells. When the virions were labeled with (3)H-leucine, purified, and adsorbed to Escherichia coli cells, about 15% of the radioactivity was recovered as low-molecular-weight material from spheroplasts formed by lysozyme-ethylenediaminetetraacetic acid. Other experiments revealed that about 7% of the total parental virus protein label could be recovered in newly formed progeny virus.
Insights
Bacteriophage phiX174 attaches to bacterial cell walls, releasing its DNA. Specific treatments reveal attachment sites and prevent DNA release, offering insights into phage-host interactions.
Area of Science:
- Microbiology
- Virology
- Molecular Biology
Background:
- Bacteriophage phiX174 is an icosahedral phage.
- It lacks a complex tail assembly for host cell attachment.
Purpose of the Study:
- To investigate the attachment mechanism of bacteriophage phiX174 to bacterial host cells.
- To understand the role of cell wall components and phage structure in DNA release.
Main Methods:
- Incubation of phiX174 virions with isolated bacterial cell walls.
- Treatment of cell walls with chloroform, lysozyme, and virions with formaldehyde.
- Freeze-etch electron microscopy of adsorbed phages on Escherichia coli.
- Radioactive labeling of phage proteins and recovery from spheroplasts.
Main Results:
- Virions attached to cell wall fragments, leading to phage deoxyribonucleic acid (DNA) release.
- Chloroform treatment prevented attachment; lysozyme or formaldehyde treatment prevented DNA release but not attachment.
- Lysozyme treatment released structures that bound phiX174 at vertices, but without DNA release.
- Phages adsorbed to bacteria were submerged in the cell wall with their fivefold axis normal to the surface.
- Phage proteins were recovered as low-molecular-weight material and in progeny viruses.
Conclusions:
- Bacteriophage phiX174 attachment involves specific interactions with bacterial cell wall components.
- DNA release is a distinct step following attachment, dependent on cell wall integrity and phage viability.
- The phage's fivefold axis of symmetry plays a role in its orientation and interaction with the host cell surface.