Replication of bacteriophage MS2 X. Phage-specific Ribonucleoprotein Particles found in MS2-infected Escherichia coli

Abstract

Phage-specific RNA-protein complexes formed during the MS2 infection process were examined. The fate of 32P-labeled parental viral RNA was followed to determine what RNA-protein interactions developed early in infection. In order to identify phage-specific ribonucleoprotein complexes at later times in infection, their protein or RNA components were labeled selectively with radioisotopes after suppression of bacterial macromolecular syntheses with Miracil D (Burroughs Wellcome and Co.). During the first nine minutes of infection, parental RNA was found in three phage-specific complexes: a “replicative complex” containing RNA synthetase and replicative intermediate RNA which sedimented faster than the 50 s ribosomal subunit; a particle composed of viral synthetase and single-stranded viral RNA sedimenting at approximately 40 s; and single-stranded viral RNA attached to a ribosomal subunit, also sedimenting at approximately 40 s. The only parental RNA not bound to protein or ribosomal subunits had been degraded. Later in infection, ribonucleoprotein particles similar to those found at early times, plus intermediates in the assembly of viral particles, were present. The replicative complex, as well as viral assembly intermediates, sedimented between 50 and 81 s. Three phage-specific species sedimented in a composite peak at approximately 45 s: a synthetase-viral RNA complex; viral RNA attached to a 30 s ribosomal subunit (both observed at earlier times of infection); plus a third species containing single-stranded progeny RNA, viral coat protein, and viral maturation protein. This third species appeared to be a direct precursor of mature phage particles.