Laura K. Parker

Bacteriophages play an important role in the pathogenicity of Staphylococcus aureus , an important human pathogen. Phages are involved in generalized and specialized transduction as well as a more specific process by which they mobilize elements known as phage-inducible chromosomal islands, of which S. aureus pathogenicity islands (SaPIs) are an important group. SaPIs are mobilized at high frequency through interactions with specific “helper” bacteriophages, such as 80α, leading to packaging of the SaPI genomes into virions made from structural proteins supplied by the helper. Among these structural proteins is the portal protein, which forms a ring-like portal at a fivefold vertex of the capsid, through which the DNA is packaged during virion assembly and ejected upon infection of the host. We previously showed that portal protein expressed in E. coli forms tridecameric rings, while portals found in virions are always dodecamers. To understand the role of the portal in capsid assembly, DNA packaging and ejection, we have here examined this phenomenon further. We show that portals assembled at lower temperature form unclosed rings that may represent portal assembly intermediates. By analyzing portal protein deletion mutants, we demonstrate the involvement of the different functional domains in phage assembly and protein incorporation.

Tailed bacteriophages with double-stranded DNA genomes (class Caudoviricetes ) play an important role in the evolution of bacterial pathogenicity, both as carriers of genes encoding virulence factors and as the main means of horizontal transfer of mobile genetic elements (MGEs) in many bacteria, such as Staphylococcus aureus . The S. aureus pathogenicity islands (SaPIs), including SaPI1, are a type of MGEs are that carry a variable complement of genes encoding virulence factors. SaPI1 is mobilized at high frequency by “helper” bacteriophages, such as 80α, leading to packaging of the SaPI1 genome into virions made from structural proteins supplied by the helper. 80α and SaPI1 virions consist of an icosahedral head (capsid) connected via a unique vertex to a long, non-contractile tail. At one end of the tail, proteins associated with the baseplate recognize and bind to the host. At the other end, a connector or “neck” forms the interface between the tail and the head. The neck consists of several specialized proteins with specific roles in DNA packaging, phage assembly, and DNA ejection. Using cryo-electron microscopy and three-dimensional reconstruction, we have determined the high-resolution structure of the neck section of SaPI1 virions made in the presence of phage 80α, including the dodecameric portal (80α gene product (gp) 42) and head-tail-connector (gp49) proteins, the hexameric head-tail joining (gp50) and tail terminator (gp52) proteins, and the major tail protein (gp53) itself. We were also able to resolve the DNA, the tail completion protein (gp51) and the tape measure protein (gp56) inside the tail. This is the first detailed structural description of these features in a bacteriophage, providing insights into the assembly and infection process in this important group of MGEs and their helper bacteriophages.