Characterization of two abundant mRNAs of Autographa californica nuclear polyhedrosis virus present late in infection

Abstract

Cytoplasmic poly(A) + RNA isolated from Spodoptera frugiperda cells late after infection with Autographa californica nuclear polyhedrosis virus (30–40 hr pi) was fractionated according to size on denaturing methyl mercury gels. Two major RNA species (1.4 kb and 0.75 kb) and several minor RNA species were detected by ethidium bromide staining. The predominant RNA species of about 1.4 kb was considered to be polyhedrin mRNA because (1) in vitro translation of the RNA, which was eluted from methyl mercury gels, yielded a polypeptide of MW 33K, which comigrated with polyhedrin. (2) When poly(A) + RNA was fractionated on a sucrose column and then translated in vitro, the distribution and abundance profiles of a 33K polypeptide product and of 1.4-kb RNA were similar. (3) The 33K polypeptide made in vitro and purified polyhedrin gave rise to similar patterns of peptides when digested with S. aureus V8 protease. The polyhedrin mRNA (1.4 kb) hybridized to BamHI-F and HindIII-V AcNPV DNA fragments and hybridization selection with BamHI-F AcNPV DNA yielded a 33K polypeptide, which comigrated with polyhedrin. The second RNA species (0.75 kb in size) hybridized to overlapping EcoRI-P and HindIII-Q regions of the AcNPV genome and translated into a methionine deficient polypeptide of MW = 8K. It was synthesized in large quantities late in the infection and appeared to be coordinately expressed with polyhedrin in infected cells. The 8K polypeptide was detected as early as 15 hr pi and was still synthesized at 60 hr pi.