CHARACTERIZATION OF BACULOVIRUS EARLY GENE EXPRESSION

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The baculoviruses are a family of large DNA viruses pathogenic to insects. They are currently employed as important biological controls of insect pests and are being engineered for improved toxicity and wider host ranges. Evaluations on the safety of these viruses in regard to non-target organisms, including humans, have demanded an increased understanding of their molecular biology and the mechanisms which govern their replication. From a virological perspective, the baculoviruses are of interest because of their large, superhelical genome, their complex replicative cycle resulting in an unusual production of two infectious forms of progeny, and the ability of the viral genome to incorporate host- derived transposable elements. We propose to examine the molecular mechanisms involved in the expression of the earliest of viral genes using the model baculovirus, Autographa californica nuclear polyhedrosis virus (AcNPV). Our objectives include defining the cis-acting DNA sequences which mediate immediate- early (IE) transcription employing both transient expression of chimeric genes and expression of such genes after their transfer back into the viral genome. We will characterize the products of two adjacent IE genes and examine their potential regulation by overlapping, anti-sense RNAs. A major portion of this proposal also examines the nature and mutagenic effects of a newly identified transposable element (TED) inserted within an IE region of the AcNPV genome. TED is a retrotransposon which upon integration directs abundant transcription of neighboring viral genes. Retrotransposons closely resemble the retroproviruses and represent an important class of insertion mutagens in eukaryotes. The exact mechanisms of their mutagenesis are unknown. We will analyze in detail the effects of TED integration on the expression of AcNPV genes. In addition, we will take advantage of the clonal isolate of TED within the viral genome to examine the structural organization and regulation of expression of a transcriptionally active retrotransposon. These studies are expected to yield further insight into the mechanisms of genetic alterations caused by these mobile elements as well as information on the complex mechanisms which control eukaryotic gene expression.