Dissimilar expression of Autographa californica multiple nucleocapsid nuclear polyhedrosis virus polyhedrin and p10 genes

Abstract

The temporal expression of the Autographa californica multiple nucleocapsid nuclear polyhedrosis virus polyhedrin and p10 genes in Spodoptera frugiperda cells was studied using virus recombinants in which either gene was replaced by the juvenile hormone esterase (JHE) gene of Heliothis virescens. The JHE served as a highly specific and sensitive reporter for gene expression. Activation of the p10 gene followed a pattern different to that of polyhedrin. The p10 gene was activated a few hours earlier than the polyhedrin gene, but its expression reached a lower maximum level. Northern blot analysis complemented and confirmed the results obtained from the JHE assays. Co-infection of sense recombinants and those containing an antisense copy of the JHE gene in place of the polyhedrin or p10 gene resulted in reduced levels of JHE gene expression. These experiments independently supported the hypothesis that the p10 gene promoter is more active at earlier times post-infection than that of the polyhedrin gene. The results also highlight the potential of the antisense strategy as an experimental approach for the study of baculovirus gene regulation and possibly insect metabolism.

Superior expression of juvenile hormone esterase and beta-galactosidase from the basic protein promoter of Autographa californica nuclear polyhedrosis virus compared to the p10 protein and polyhedrin promoters

“…In the first of these studies expression of JHE under control of the pl0 promoter was consistently higher than expression under control of the polyhedrin promoter. In contrast, Roelvink et al (1992) found that the reverse was true for the pl0-negative virus at later times after infection.…”

“…The coding sequence includes a 19 amino acid leader which signals export of expressed JHE from cells into the medium in cell culture. In vitro analysis of expression of JHE by recombinant viruses showed that the pl0 promoter is active several hours earlier than the polyhedrin promoter Roelvink et al, 1992). Conflicting results concerning the relative yields of JHE expressed under the control of the pl0 promoter in pl0-positive ) and pl0-negative (Roelvink et aL, 1992) viruses prompted further comparative analysis of these expression vectors.…”

“…In vitro analysis of expression of JHE by recombinant viruses showed that the pl0 promoter is active several hours earlier than the polyhedrin promoter Roelvink et al, 1992). Conflicting results concerning the relative yields of JHE expressed under the control of the pl0 promoter in pl0-positive ) and pl0-negative (Roelvink et aL, 1992) viruses prompted further comparative analysis of these expression vectors. In the first of these studies expression of JHE under control of the pl0 promoter was consistently higher than expression under control of the polyhedrin promoter.…”

Gene gymnastics

“…12,23,24 This is exacerbated for the viruses of the BAC/Tn7 type by the fact that the insertion site which is targeted by the Tn7 transposon is actually a mutational hotspot. 29,30 It is clear that the possibilities to improve the baculovirus itself are ample, and are far from exhausted in state of the art systems.…”

Lepidopteran cells, an alternative for the production of recombinant antibodies?

“…16 P10 gene is expressed a few hours before polyhedrin, thus leading to the overexpression of the H chains before the light chains and triggering of secretion of H chains dimers (H 2 ). 15 Since the correct folding of the heavy chain requires the presence of light chains, the unfolded H2 molecules may be at the origin of the formation of aggregates observed under these conditions. 17 Aggregation may in turn also explain the low expression rate observed in these conditions.…”