Control of membrane morphogenesis in bacteriophage PM2

Abstract

The regulation of membrane formation in bacteriophage PM2 serves as a simple model for changes in membrane structure in eukaryotic cells. Prior to Pseudomonas host lysis, wild-type virions mature to an icosahedral morphology at the inner face of the cytoplasmic membrane. The preliminary characterization of two temperature-sensitive mutants of PM2 is described. In cells infected at the restrictive temperature with ts 1, an abundance of "empty" virus-size membrane vesicles are seen. Synthesis of DNA is also reduced in ts 1 infected cells. The preponderance of vesicles is not seen in cells infected with wild-type virus or with ts 1 at the permissive temperature. The "empty" appearance of the viral membranes suggests that viral DNA is not encapsulated. The major viral capsid protein (MW 26,000) is located just outside the viral membrane and normally sidiments with host and virus membranes. This protein made by mutant ts 5 does not pellet with these membranes; instead, large amounts of capsid protein can be precipitated from the supernatant with TCA. Compared to cells infected with wild type virus, cells infected with ts 5 at the restrictive temperature produced inside the cell an abundance of virus-size membrane vesicles. Taken together, these results with viral mutants suggest that formation of a viral membrane of the proper size does not require a DNA core around which to form, or an outer scaffolding of coat protein against which to form a spherical bilayer.

Bacteriophage PM2 Has a Protein Capsid Surrounding a Spherical Proteinaceous Lipid Core

“…P1, with a molecular mass of 37.5 kDa, has been identified as the spike protein located at the fivefold vertices, and P2, with a molecular mass of 30.2 kDa, is the major capsid protein. Proteins P3 and P4 are located internally (9,10,22,36,37,38,41,45,46), and P4 has been proposed to be an integral membrane protein interacting with the viral DNA (9,40,42). Dissociation of the PM2 particle has been extensively studied.…”

Mutant ts1 of bacteriophage PM2 is defective in the major capsid protein and fails to package its DNA

“…Alternatively, the DNA may have been packaged, but upon manipulation or isolation of the vesicles, it was released from inside the membrane. Although precedent exists in the case of 10-and 26mutants of bacteriophage P22 (3), this explanation seems less likely, since the vesicles of sectioned cells appear empty (4). The intermediate state of a linear molecule could be tested by exposing cell-free extracts of infected cells (16) to purified wild-type sp27.…”

Assembly of bacteriophage PRD1: particle formation with wild-type and mutant viruses

“…P. aeruginosa was used for these experiments because of the ease and completeness of cell lysis. Cells were infected at a multiplicity of infection of 20, and [35S]methionine (5 pCi/ml) was added at 27 min postinfection. Labeling was terminated 2 min later by the addition of 50 ,ug of L-methionine per ml.…”