Developmental variation in cytochrome P450 expression in Papilio polyxenes in response to xanthotoxin, a hostplant allelochemical

Terry L. Harrison, Arthur R. Zangerl, Mary A. Schuler, May R. Berenbaum

Research output: Contribution to journalArticle

Abstract

Although developmental variation in activity and inducibility is typical of cytochrome P450 monooxygenases (P450s) in insects, the adaptive significance of such variation is often unclear, in part because the natural function of insect P450s is rarely known. In this study, we examined developmental variation in expression of CYP6B1 and CYP6B3 in Papilio polyxenes, the black swallowtail. Enzymes encoded by these genes have been implicated in the metabolism of xanthotoxin, a furanocoumarin characteristic of the apiaceous hostplants of P. polyxenes. In each life stage - egg, five larval instars, pupa, and adult - we examined individuals exposed to foliage with and without supplemental xanthotoxin. For each stage, we conducted enzyme assays to estimate xanthotoxin metabolism, Northern analysis to detect constitutive and induced mRNA levels, and RT-PCR amplification and Southern analysis to differentiate among P450 genes expressed. Inducible xanthotoxin metabolism, previously reported in fifth instars, was observed in four of five larval stages but was absent or undetectable in all stages that do not feed on foliage; the highest levels of activity were in early larval instars. The same pattern was observed in both Northern and RT-PCR gel blot analyses. In inducible larval stages, inducibility of CYP6B1 transcripts by xanthotoxin was greater than the inducibility of CYP6B3 transcripts. These findings support earlier suggestions that these two P450s contribute to xanthotoxin metabolism in this species and that expression of these P450 genes is regulated in an adaptive fashion with respect to probability of exposure to hostplant toxins over the course of development.

Original languageEnglish (US)
Pages (from-to)179-189
Number of pages11
JournalArchives of Insect Biochemistry and Physiology
Volume48
Issue number4
DOIs
StatePublished - Dec 1 2001

Fingerprint

Papilio polyxenes
methoxsalen
Methoxsalen
Pheromones
allelochemicals
cytochrome P-450
Cytochrome P-450 Enzyme System
Metabolism
Mixed Function Oxygenases
metabolism
instars
Genes
Insects
reverse transcriptase polymerase chain reaction
psoralens
Polymerase Chain Reaction
Pupa
insects
genes
Enzyme Assays

Keywords

  • Cytochrome P450
  • Furanocoumarins
  • Papilio polyxenes
  • Transcriptional regulation

ASJC Scopus subject areas

  • Physiology
  • Biochemistry
  • Insect Science

Cite this

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title = "Developmental variation in cytochrome P450 expression in Papilio polyxenes in response to xanthotoxin, a hostplant allelochemical",
abstract = "Although developmental variation in activity and inducibility is typical of cytochrome P450 monooxygenases (P450s) in insects, the adaptive significance of such variation is often unclear, in part because the natural function of insect P450s is rarely known. In this study, we examined developmental variation in expression of CYP6B1 and CYP6B3 in Papilio polyxenes, the black swallowtail. Enzymes encoded by these genes have been implicated in the metabolism of xanthotoxin, a furanocoumarin characteristic of the apiaceous hostplants of P. polyxenes. In each life stage - egg, five larval instars, pupa, and adult - we examined individuals exposed to foliage with and without supplemental xanthotoxin. For each stage, we conducted enzyme assays to estimate xanthotoxin metabolism, Northern analysis to detect constitutive and induced mRNA levels, and RT-PCR amplification and Southern analysis to differentiate among P450 genes expressed. Inducible xanthotoxin metabolism, previously reported in fifth instars, was observed in four of five larval stages but was absent or undetectable in all stages that do not feed on foliage; the highest levels of activity were in early larval instars. The same pattern was observed in both Northern and RT-PCR gel blot analyses. In inducible larval stages, inducibility of CYP6B1 transcripts by xanthotoxin was greater than the inducibility of CYP6B3 transcripts. These findings support earlier suggestions that these two P450s contribute to xanthotoxin metabolism in this species and that expression of these P450 genes is regulated in an adaptive fashion with respect to probability of exposure to hostplant toxins over the course of development.",
keywords = "Cytochrome P450, Furanocoumarins, Papilio polyxenes, Transcriptional regulation",
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T1 - Developmental variation in cytochrome P450 expression in Papilio polyxenes in response to xanthotoxin, a hostplant allelochemical

AU - Harrison, Terry L.

AU - Zangerl, Arthur R.

AU - Schuler, Mary A.

AU - Berenbaum, May R.

PY - 2001/12/1

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N2 - Although developmental variation in activity and inducibility is typical of cytochrome P450 monooxygenases (P450s) in insects, the adaptive significance of such variation is often unclear, in part because the natural function of insect P450s is rarely known. In this study, we examined developmental variation in expression of CYP6B1 and CYP6B3 in Papilio polyxenes, the black swallowtail. Enzymes encoded by these genes have been implicated in the metabolism of xanthotoxin, a furanocoumarin characteristic of the apiaceous hostplants of P. polyxenes. In each life stage - egg, five larval instars, pupa, and adult - we examined individuals exposed to foliage with and without supplemental xanthotoxin. For each stage, we conducted enzyme assays to estimate xanthotoxin metabolism, Northern analysis to detect constitutive and induced mRNA levels, and RT-PCR amplification and Southern analysis to differentiate among P450 genes expressed. Inducible xanthotoxin metabolism, previously reported in fifth instars, was observed in four of five larval stages but was absent or undetectable in all stages that do not feed on foliage; the highest levels of activity were in early larval instars. The same pattern was observed in both Northern and RT-PCR gel blot analyses. In inducible larval stages, inducibility of CYP6B1 transcripts by xanthotoxin was greater than the inducibility of CYP6B3 transcripts. These findings support earlier suggestions that these two P450s contribute to xanthotoxin metabolism in this species and that expression of these P450 genes is regulated in an adaptive fashion with respect to probability of exposure to hostplant toxins over the course of development.

AB - Although developmental variation in activity and inducibility is typical of cytochrome P450 monooxygenases (P450s) in insects, the adaptive significance of such variation is often unclear, in part because the natural function of insect P450s is rarely known. In this study, we examined developmental variation in expression of CYP6B1 and CYP6B3 in Papilio polyxenes, the black swallowtail. Enzymes encoded by these genes have been implicated in the metabolism of xanthotoxin, a furanocoumarin characteristic of the apiaceous hostplants of P. polyxenes. In each life stage - egg, five larval instars, pupa, and adult - we examined individuals exposed to foliage with and without supplemental xanthotoxin. For each stage, we conducted enzyme assays to estimate xanthotoxin metabolism, Northern analysis to detect constitutive and induced mRNA levels, and RT-PCR amplification and Southern analysis to differentiate among P450 genes expressed. Inducible xanthotoxin metabolism, previously reported in fifth instars, was observed in four of five larval stages but was absent or undetectable in all stages that do not feed on foliage; the highest levels of activity were in early larval instars. The same pattern was observed in both Northern and RT-PCR gel blot analyses. In inducible larval stages, inducibility of CYP6B1 transcripts by xanthotoxin was greater than the inducibility of CYP6B3 transcripts. These findings support earlier suggestions that these two P450s contribute to xanthotoxin metabolism in this species and that expression of these P450 genes is regulated in an adaptive fashion with respect to probability of exposure to hostplant toxins over the course of development.

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