This research proposal focuses on phasmid P4, a replicon that can be propagated in most gram negative bacteria in any of three modes: (i) as a lytic virus dependent on a helper phage such as P2; (ii) as a prophage, or (iii) as a plasmid. The long term objectives of our studies with P4 are to characterize fundamental molecular mechanisms by which: (i) the sizes of icosahedral virion capsids are determined, and (ii) the genes of bacteria and bacterial viruses are regulated. At the morphogenic level the specific aims of this proposal are: (i) to resolve the basic steps in capsid assembly that are specified by helper virus P2, and (ii) to decipher the targeted assembly step in the P2 morphogenic pathway at which the P4-encoded sid (size determination) gene product acts and the associated mechanism of action. This will include testing the specific hypothesis that sid acts as a scaffolding-type morphogenic protein. This will be tested based on the inducible expression of the sid gene from a cloning vector during lytic infection by the helper, and during expression of morphogenic genes of the helper virus that that have been cloned for this purpose. At the level of gene regulation the specific aims of this proposal are: (i) to test the generality of the P4-encoded psu gene product as a transcription antitermination factor and to decipher both the mechanistic level and targets involved, and (ii) to directly test the hypothesis that the P4-encoded sid gene product likewise acts as an antiterminator, and if so, to determine how such activity plays a role in capsid size determination. For these studies, psu and sid proteins will each be inducibly expressed from cloning vectors and their activities monitored with respect to effecting transcriptional patterns of the P2 genome and also that of well characterized bacterial operons that have been cloned for this purpose. The health relatedness of this project will be derived from fundamental insights into the molecular mechanisms by which viruses and bacteria are propagated.
| Sajjan, U S; Sun, L; Goldstein, R et al. (1995) Cable (cbl) type II pili of cystic fibrosis-associated Burkholderia (Pseudomonas) cepacia: nucleotide sequence of the cblA major subunit pilin gene and novel morphology of the assembled appendage fibers. J Bacteriol 177:1030-8 |
