title: An RNA Domain Imparts Specificity and Selectivity to a Viral DNA Packaging Motor.
abstract: During assembly, double-stranded DNA viruses, including bacteriophages and herpesviruses, utilize a powerful molecular motor to package their genomic DNA into a preformed viral capsid. An integral component of the packaging motor in the Bacillus subtilis bacteriophage ϕ29 is a viral genome-encoded pentameric ring of RNA (prohead RNA [pRNA]). pRNA is a 174-base transcript comprised of two domains, domains I and II. Early studies initially isolated a 120-base form (domain I only) that retains high biological activity in vitro; hence, no function could be assigned to domain II. Here we define a role for this domain in the packaging process. DNA packaging using restriction digests of ϕ29 DNA showed that motors with the 174-base pRNA supported the correct polarity of DNA packaging, selectively packaging the DNA left end. In contrast, motors containing the 120-base pRNA had compromised specificity, packaging both left- and right-end fragments. The presence of domain II also provides selectivity in competition assays with genomes from related phages. Furthermore, motors with the 174-base pRNA were restrictive, in that they packaged only one DNA fragment into the head, whereas motors with the 120-base pRNA packaged several fragments into the head, indicating multiple initiation events. These results show that domain II imparts specificity and stringency to the motor during the packaging initiation events that precede DNA translocation. Heteromeric rings of pRNA demonstrated that one or two copies of domain II were sufficient to impart this selectivity/stringency. Although ϕ29 differs from other double-stranded DNA phages in having an RNA motor component, the function provided by pRNA is carried on the motor protein components in other phages.
  
  
  OBJECTIVE
  During virus assembly, genome packaging involves the delivery of newly synthesized viral nucleic acid into a protein shell. In the double-stranded DNA phages and herpesviruses, this is accomplished by a powerful molecular motor that translocates the viral DNA into a preformed viral shell. A key event in DNA packaging is recognition of the viral DNA among other nucleic acids in the host cell. Commonly, a DNA-binding protein mediates the interaction of viral DNA with the motor/head shell. Here we show that for the bacteriophage ϕ29, this essential step of genome recognition is mediated by a viral genome-encoded RNA rather than a protein. A domain of the prohead RNA (pRNA) imparts specificity and stringency to the motor by ensuring the correct orientation of DNA packaging and restricting initiation to a single event. Since this assembly step is unique to the virus, DNA packaging is a novel target for the development of antiviral drugs.
authors.affiliation: Department of Diagnostic and Biological Sciences and Institute for Molecular Virology, University of Minnesota, Minneapolis, Minnesota, USA.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhao
authors.affiliation: Department of Diagnostic and Biological Sciences and Institute for Molecular Virology, University of Minnesota, Minneapolis, Minnesota, USA.
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Jardine
authors.affiliation: Department of Diagnostic and Biological Sciences and Institute for Molecular Virology, University of Minnesota, Minneapolis, Minnesota, USA grime001@umn.edu.
authors.forename: Shelley
authors.initials: S
authors.lastname: Grimes
mesh_terms: D017103:Bacillus Phages / Q000502:physiology* / Q000648:ultrastructure; D004279:DNA, Viral / Q000235:genetics / Q000378:metabolism*; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D019065:Virus Assembly / Q000502:physiology*
chemical_list: D004279:DNA, Viral; D012367:RNA, Viral; D014764:Viral Proteins