Comparative analysis of the transcripts mapped in the BamHI DNA fragment B of avirulent HSV-1 HFEM, virulent HSV-1 F, and their intratypic recombinant viruses

Abstract

HSV-1 HFEM, whose genome harbors a deletion of 4.1 kbp (0.762 to 0.789 map units (mu] is avirulent for mice and tree shrews by the intraperitoneal (i.p.) application route. Insertion of the BamHI DNA fragment B (0.738 to 0.809 mu) and/or the MluI DNA fragment (0.7615 to 0.796 mu) molecularly cloned from virulent HSV-1 F, restored the i.p. pathogenicity to strain HFEM and led to the isolation of virulent intratypic recombinants. In order to determine the RNA transcripts mapped in the BamHI DNA fragment B of the HSV-1 HFEM, HSV-1 F, and their intratypic recombinants R15, R19, R26, and R-Ml-C1, a comparative analysis was performed using Northern blot hybridizations. Two novel RNA transcripts of 3.5 and 1.5 kb were detected which hybridize to the left terminus (0.738 to 0.746 mu) of the BamHI DNA fragment B. The 1.5 kb RNA transcript was missing in the avirulent HSV-1 HFEM. Hybridization with the BssHII DNA fragment F (0.760 to 0.762 mu) led to detection of a 3.5 kb RNA transcript by HSV-1 HFEM which was missing in all other viruses tested. In contrast a 1.5 kb RNA transcript was detectable in all other virus strains with the exception of HSV-1 HFEM. The 3.5 kb transcript hybridized to the right-hand flank of the deleted region in the genome of HSV-1 HFEM (Asp718/SalI DNA fragment; 0.786 to 0.79 mu). The detection of the novel 1.5 kb RNA, which is missing in HSV-1 HFEM, and the appearance of the newly transcribed 3.5 kb RNA in HSV-1 HFEM only, indicates a new open reading frame in this particular region as a consequence of the fusion of the DNA sequences at both ends of the deletion in the genome of HSV-1 HFEM.