HEADFUL PACKAGING REVISITED - THE PACKAGING OF MORE THAN ONE DNA MOLECULE INTO A BACTERIOPHAGE-P1 HEAD

Like a variety of other bacteriophages, such as T4 and P22, bacterioph age P1 packages DNA by a ''headful'' mechanism in which the capacity o f the viral capsid determines the size of the single DNA molecule that is packaged. Because of the long-standing and general acceptance of t his packaging mechanism, we were surprised to discover that some of ou r observations, using the in vitro P1 packaging system, could be expla ined by the packaging of less than headful-sized (<110 kb) DNA molecul es into a PI capsid. To account for these observations, we describe re sults that support a model of in vitro P1 packaging in which multiple less than headful-sized DNA molecules are taken into a P1 head until t hat head has been filled. The results further suggest that the phage s o generated can occasionally inject more than one DNA molecule into a cell upon viral infection. The data that supports these conclusions ar e: (1) the DNAs of the circular P1 cloning vectors pAd10sacBII (32 kb) and pNS358 (14 kb) are packaged in vitro with an efficiency of about 6 to 12 % of that of longer concatemers of these DNAs. (2) The in vitr o packaging of two differentially marked, less than 18 kb plasmid DNAs in the same reaction results in the production of a phage that can oc casionally inject both DNAs into the same cell upon infection. (3) Vir us particles generated by the packaging of either pAd10sacBII plasmid DNA or the two differently marked plasmids have a density in CsCl equi librium gradients that is the same as P1 plaque-forming phage, suggest ing that the former phage contain a headful of DNA. These results cann ot be explained by Cre-mediated site-specific recombination between pl asmids in the P1 packaging extracts. Finally we present in vivo experi ments that are also consistent with the headful packaging of multiple DNAs into a P1 head.