A novel gene in the takeout gene family is regulated by hormones and nutrients in Manduca larval epidermis.

A novel gene, moling, was cloned from epidermal RNA of the tobacco hornworm, Manduca sexta, using PCR-based suppression subtractive hybridization. moling belongs to a gene family that includes several lepidopteran hemolymph juvenile hormone (JH) binding proteins and takeout of Drosophila melanogaster. The mRNA first appears in the epidermis on day 0 of the fifth instar and rises to its peak expression by mid-day 2, then declines rapidly and is gone by the onset of wandering. moling is expressed exclusively in the last instar larval epidermis and not in the imaginal discs or any other tissues. Allatectomy early in the fourth instar induces precocious metamorphosis and causes the appearance of moling mRNA by 33 h. Allatectomy after the critical period for JH in the final larval molt had no effect on the timing of the onset of moling expression in the final instar but caused a more rapid up-regulation once begun. The JH mimic pyriproxifen given at the outset of the final instar suppressed the expression of moling mRNA to low levels, in both intact and allatectomized larvae. Starvation immediately after ecdysis to the fifth instar prevented the onset of expression. Thus, initiation of transcription requires both nutrient intake and decline in JH. Infusion of 20-hydroxyecdysone (20E) into ligated abdomens of day 2 fifth instar larvae and culture of the day 2 fifth instar larval abdominal epidermis with 20E in vitro both caused a rapid decline of moling mRNA. The slower and variable decline that occurred in mid-day 2 fifth instar larval epidermis in the ligated abdomens or when incubated in hormone-free medium indicated that the increase of 20E on day 2 had already initiated the decline of expression. The role of Moling may be to stabilize JH in the epidermal cell during the final intermolt when the JH esterase activity increases.