A cytochrome P450 terpenoid hydroxylase linked to the suppression of insect juvenile hormone synthesis

Abstract

A cDNA encoding a cytochrome P450 enzyme was isolated from a cDNA library of the corpora allata (CA) from reproductively active Diploptera punctata cockroaches. This P450 from the endocrine glands that produce the insect juvenile hormone (JH) is most closely related to P450 proteins of family 4 and was named CYP4C7. The CYP4C7 gene is expressed selectively in the CA; its message could not be detected in the fat body, corpora cardiaca, or brain, but trace levels of expression were found in the midgut and caeca. The levels of CYP4C7 mRNA in the CA, measured by ribonuclease protection assays, were linked to the activity cycle of the glands. In adult females, CYP4C7 expression increased immediately after the peak of JH synthesis, reaching a maximum on day 7, just before oviposition. mRNA levels then declined after oviposition and during pregnancy. The CYP4C7 protein was produced in Escherichia coli as a C-terminal His-tagged recombinant protein. In a reconstituted system with insect NADPH cytochrome P450 reductase, cytochrome b 5 , and NADPH, the purified CYP4C7 metabolized (2E,6E)-farnesol to a more polar product that was identified by GC-MS and by NMR as (10E)-12-hydroxyfarnesol. CYP4C7 converted JH III to 12-trans-hydroxy JH III and metabolized other JH-like sesquiterpenoids as well. This -hydroxylation of sesquiterpenoids appears to be a metabolic pathway in the corpora allata that may play a role in the suppression of JH biosynthesis at the end of the gonotrophic cycle.Juvenile hormone (JH) plays a central role in insect development, metamorphosis, and reproduction. This sesquiterpenoid epoxide is synthesized in endocrine glands, the corpora allata (CA)(1), and is degraded predominantly by esterases and epoxide hydrolases (2). The rate of JH synthesis by the CA is a major determinant of the titer of JH in the hemolymph (3), and the regulation of JH synthesis is seen as a potential target for insect control. The biosynthesis of JH has been extensively characterized during the reproductive cycle of the cockroach Diploptera punctata, an insect that serves as a convenient model system. In adult females of this insect, the cycle of JH synthesis is regulated by humoral factors and by innervation from neurosecretory cells in the brain (1). Production of JH by the CA increases 10-fold to reach a peak 5 days after adult emergence and mating, and this peak corresponds to the peak of vitellogenesis. Synthesis is then rapidly repressed and remains low from deposition of the eggs into the brood sac through pregnancy (4).The allatostatins, a family of brain-gut peptides (5-9) are known to inhibit JH synthesis in D. punctata at the end of a gonotrophic cycle, but these peptides are probably not alone responsible for repression of the intrinsic rate of JH synthesis in postvitellogenic insects. The mechanism of stable suppression of JH synthesis at the end of the cycle has received very little attention, in part because of the absence of molecular tools to dissect it. This repression occurs concomitant with changes in c...

BAS1 : A gene regulating brassinosteroid levels and light responsiveness in Arabidopsis

“…Though ecdysteroids are inactivated in a similar manner to BRs, these insect hormones do not induce BR responses in plants (39). Other insect juvenile hormones can be hydroxylated and presumably catabolized in a similar manner to ecdysone (40). CYP450-dependent C24-hydroxylation of 1,25-dihydroxyvitamin D-3 inactivates this form of vitamin D in both rats (41,42) and humans (43).…”

Vitellogenesis and Post-Vitellogenic Maturation of the Insect Ovarian Follicle

“…In the cockroach, Diploptera punctata, termination of vitellogenesis can be associated with a decline in JH titer (Sutherland et al, 1998). Ovarian follicles that are in different stages of oogenesis exert different effects on the activity of the CA.…”

“…Thus, follicles in active vitellogenesis stimulate JH production, while those near maturity inhibit it (Sutherland et al, 2000). During the progression of vitellogenesis, the target for the ovarian inhibitory hormonal factor in the cells of the CA is proposed to be the CYP4C7 cytochrome P450 enzyme, which metabolizes JH and JH precursors by b-hydroxylation (Sutherland et al, 1998(Sutherland et al, , 2000. Thus, the decline in JH titer that is necessary to initiate post-vitellogenic events in Diploptera, is mediated through a feedback signal from the ovary to the CA, which represses JH synthesis.…”

Juvenile Hormone Diol Kinase

“…7) (4,5), suggesting that JHE is more than an inactivating enzyme. Recently, a cytochrome P450 terpenoid hydroxylase was discovered in the corpora allata of Diploptera punctata; this enzyme may play a role in the temporal suppression of JH titer in the cockroach by metabolizing JH III to 12-hydroxy-JH III (6).…”