Evidence for two juvenile hormone esterase-related genes in the Colorado potato beetle

Juvenile hormone esterase (JHE) activity in the haemolymph of the Colorado potato beetle is necessary to initiate pupation in larvae as well as diapause in adults. The enzyme appears in the haemolymph as a dimer consisting of two 57 kDa subunits. The sequence of an encoding cDNA, JHE.A, is distinct from lepidopteran JHEs. In this study, RT-PCR using primers designed on the basis of the 5'- and 3'-ends of the coding region revealed the existence of a related gene, JHE.B. The presence of two JHE-related genes was also shown by PCR amplification on genomic DNA from different individual beetles followed by restriction enzyme analysis. Both forms, probably paralogues, were transcribed since they could be amplified on messenger RNA from fat bodies. The size of the PCR products generated with mRNA and genomic DNA were both 1.6 kb, suggesting the absence of introns in the genomic JHE coding sequence. The sequence of a genomic clone, which encoded JHE.B, was 77% identical and 82% similar in amino acids compared to JHE.A. No introns were found in the coding sequence of these coleopteran JHE-related genes, in contrast to lepidopteran JHE genes. Southern blot analysis of digested genomic DNA confirmed the presence of two JHE-related genes.