Functional conservation of the wingless-engrailed interaction as shown by a widely applicable baculovirus misexpression system.

BACKGROUND: The expression patterns of the segment polarity genes wingless and engrailed are conserved during segmentation in a variety of arthropods, suggesting that the regulatory interactions between these two genes are also evolutionarily conserved. Hypotheses derived from such comparisons of gene expression patterns are difficult to test experimentally as genetic manipulation is currently possible for only a few model organisms. RESULTS: We have developed a system, using recombinant baculoviruses, that can be applied to a wide variety of organisms to study the effects of ectopic expression of genes. As a first step, we studied the range and type of infection of several reporter viruses in the embryos of two arthropod and one vertebrate species. Using this system to express wingless, we were able to induce expression of engrailed in the anterior half of each parasegment in embryos of the fruit fly Drosophila melanogaster. Virus-mediated wingless expression also caused ectopic naked ventral cuticle formation in wild-type Drosophila larvae. In the flour beetle, Tribolium castaneum, ectopic wingless also induced engrailed expression. As in Drosophila, this expression was only detectable in the anterior half of the parasegment. CONCLUSIONS: The functional interaction between wingless and engrailed, and the establishment of cells competent to express engrailed, appears to be conserved between Drosophila and Tribolium. The data on the establishment of an engrailed-competent domain also support the idea that prepatterning by pair-rule genes is conserved between these two insects. The recombinant baculovirus technology reported here may help answer other long-standing comparative evolutionary questions.

Evidence for rolling circle replication of Autographa californica M nucleopolyhedrovirus genomic DNA.

Autographa californica M nucleopolyhedrovirus (AcMNPV) is a large ds DNA virus restricted to larval lepidopteran insect hosts. Using field inversion gel electrophoresis and digestion with a restriction enzyme which cuts the AcMNPV genome once, we detected multiple unit-length genome fragments from replicating viral DNA. Our data suggest that AcMNPV replicates in a head-to-tail manner via rolling circle replication.

Proteolysis of p6.9 induced by cytochalasin D in Autographa californica M nuclear polyhedrosis virus-infected cells.

Cytochalasin D, a fungus-derived compound that interferes with actin polymerization, inhibits Autographa californica M nuclear polyhedrosis virus production in infected Spodoptera frugiperda (IPLB-Sf-21) cells. Cytochalasin D appears to inhibit nucleocapsid morphogenesis by interfering with nucleoprotein packaging. We were interested in determining, therefore, whether the drug affected the synthesis or processing of p6.9, the major core protein involved in nucleoprotein packaging. We found that cytochalasin D had no effect on the synthesis, phosphorylation, or dephosphorylation of p6.9, but that it induced the proteolysis of p6.9, an effect which could account for the inhibition of nucleocapsid morphogenesis. We also determined that the cytochalasin D-induced proteolysis of p6.9 was reversible upon removal of the drug, even in the absence of protein synthesis.