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Olfactory proteins mediating chemical communication in the navel orangeworm moth, Amyelois transitella.

Leal WS, Ishida Y, Pelletier J, Xu W, Rayo J, Xu X, Ames JB - PLoS ONE (2009)

Bottom Line: We have cloned nine cDNAs encoding olfactory proteins from the navel orangeworm, including two pheromone-binding proteins, two general odorant-binding proteins, one chemosensory protein, one glutathione S-transferase, one antennal binding protein X, one sensory neuron membrane protein, and one odorant receptor.Of these, AtraPBP1 is highly enriched in male antennae.Fluorescence, CD and NMR studies suggest a dramatic pH-dependent conformational change, with high affinity to pheromone constituents at neutral pH and no binding at low pH.

View Article: PubMed Central - PubMed

Affiliation: Department of Entomology, University of California Davis, Davis, California, United States of America. wsleal@ucdavis.edu

ABSTRACT

Background: The navel orangeworm, Amyelois transitella Walker (Lepidoptera: Pyralidae), is the most serious insect pest of almonds and pistachios in California for which environmentally friendly alternative methods of control--like pheromone-based approaches--are highly desirable. Some constituents of the sex pheromone are unstable and could be replaced with parapheromones, which may be designed on the basis of molecular interaction of pheromones and pheromone-detecting olfactory proteins.

Methodology: By analyzing extracts from olfactory and non-olfactory tissues, we identified putative olfactory proteins, obtained their N-terminal amino acid sequences by Edman degradation, and used degenerate primers to clone the corresponding cDNAs by SMART RACE. Additionally, we used degenerate primers based on conserved sequences of known proteins to fish out other candidate olfactory genes. We expressed the gene encoding a newly identified pheromone-binding protein, which was analyzed by circular dichroism, fluorescence, and nuclear magnetic resonance, and used in a binding assay to assess affinity to pheromone components.

Conclusion: We have cloned nine cDNAs encoding olfactory proteins from the navel orangeworm, including two pheromone-binding proteins, two general odorant-binding proteins, one chemosensory protein, one glutathione S-transferase, one antennal binding protein X, one sensory neuron membrane protein, and one odorant receptor. Of these, AtraPBP1 is highly enriched in male antennae. Fluorescence, CD and NMR studies suggest a dramatic pH-dependent conformational change, with high affinity to pheromone constituents at neutral pH and no binding at low pH.

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Related in: MedlinePlus

RT-PCR analysis of a pheromone-binding protein gene.As previous results suggested that AtraPBP1 is expressed exclusively in male antennae, a more stringent RT-PCR analysis was performed and expression in other non-olfactory tissues was re-examined. AtraPBP1 was detected in male antennae (Ant), but not in legs (L), wings (W), thorax (Thx), or abdomen (Abd). A faint band, hardly seen in the figure, was observed in the original gel with female cDNA template thus suggesting basal expression in female antennae. RpL8 was used as a control gene.
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pone-0007235-g004: RT-PCR analysis of a pheromone-binding protein gene.As previous results suggested that AtraPBP1 is expressed exclusively in male antennae, a more stringent RT-PCR analysis was performed and expression in other non-olfactory tissues was re-examined. AtraPBP1 was detected in male antennae (Ant), but not in legs (L), wings (W), thorax (Thx), or abdomen (Abd). A faint band, hardly seen in the figure, was observed in the original gel with female cDNA template thus suggesting basal expression in female antennae. RpL8 was used as a control gene.

Mentions: Having observed by non-quantitative RT-PCR that AtraPBP1 gene is expressed only in male antennae, a more thorough examination of gene expression was performed. Indeed, AtraPBP1 was limited to expression in male antennae (Fig. 4), with no trace detected in non-olfactory tissues, including legs, wings, thorax, and abdomen. It is worth mentioning, however, that a faint band was observed when cDNA from female antennae was used as template thus suggesting that AtraPBP1 is highly enriched in male antennae. Consequently, it is reasonable to assume that AtraPBP1 plays male-specific role(s), such as the detection of sex pheromones.


Olfactory proteins mediating chemical communication in the navel orangeworm moth, Amyelois transitella.

Leal WS, Ishida Y, Pelletier J, Xu W, Rayo J, Xu X, Ames JB - PLoS ONE (2009)

RT-PCR analysis of a pheromone-binding protein gene.As previous results suggested that AtraPBP1 is expressed exclusively in male antennae, a more stringent RT-PCR analysis was performed and expression in other non-olfactory tissues was re-examined. AtraPBP1 was detected in male antennae (Ant), but not in legs (L), wings (W), thorax (Thx), or abdomen (Abd). A faint band, hardly seen in the figure, was observed in the original gel with female cDNA template thus suggesting basal expression in female antennae. RpL8 was used as a control gene.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2749207&req=5

pone-0007235-g004: RT-PCR analysis of a pheromone-binding protein gene.As previous results suggested that AtraPBP1 is expressed exclusively in male antennae, a more stringent RT-PCR analysis was performed and expression in other non-olfactory tissues was re-examined. AtraPBP1 was detected in male antennae (Ant), but not in legs (L), wings (W), thorax (Thx), or abdomen (Abd). A faint band, hardly seen in the figure, was observed in the original gel with female cDNA template thus suggesting basal expression in female antennae. RpL8 was used as a control gene.
Mentions: Having observed by non-quantitative RT-PCR that AtraPBP1 gene is expressed only in male antennae, a more thorough examination of gene expression was performed. Indeed, AtraPBP1 was limited to expression in male antennae (Fig. 4), with no trace detected in non-olfactory tissues, including legs, wings, thorax, and abdomen. It is worth mentioning, however, that a faint band was observed when cDNA from female antennae was used as template thus suggesting that AtraPBP1 is highly enriched in male antennae. Consequently, it is reasonable to assume that AtraPBP1 plays male-specific role(s), such as the detection of sex pheromones.

Bottom Line: We have cloned nine cDNAs encoding olfactory proteins from the navel orangeworm, including two pheromone-binding proteins, two general odorant-binding proteins, one chemosensory protein, one glutathione S-transferase, one antennal binding protein X, one sensory neuron membrane protein, and one odorant receptor.Of these, AtraPBP1 is highly enriched in male antennae.Fluorescence, CD and NMR studies suggest a dramatic pH-dependent conformational change, with high affinity to pheromone constituents at neutral pH and no binding at low pH.

View Article: PubMed Central - PubMed

Affiliation: Department of Entomology, University of California Davis, Davis, California, United States of America. wsleal@ucdavis.edu

ABSTRACT

Background: The navel orangeworm, Amyelois transitella Walker (Lepidoptera: Pyralidae), is the most serious insect pest of almonds and pistachios in California for which environmentally friendly alternative methods of control--like pheromone-based approaches--are highly desirable. Some constituents of the sex pheromone are unstable and could be replaced with parapheromones, which may be designed on the basis of molecular interaction of pheromones and pheromone-detecting olfactory proteins.

Methodology: By analyzing extracts from olfactory and non-olfactory tissues, we identified putative olfactory proteins, obtained their N-terminal amino acid sequences by Edman degradation, and used degenerate primers to clone the corresponding cDNAs by SMART RACE. Additionally, we used degenerate primers based on conserved sequences of known proteins to fish out other candidate olfactory genes. We expressed the gene encoding a newly identified pheromone-binding protein, which was analyzed by circular dichroism, fluorescence, and nuclear magnetic resonance, and used in a binding assay to assess affinity to pheromone components.

Conclusion: We have cloned nine cDNAs encoding olfactory proteins from the navel orangeworm, including two pheromone-binding proteins, two general odorant-binding proteins, one chemosensory protein, one glutathione S-transferase, one antennal binding protein X, one sensory neuron membrane protein, and one odorant receptor. Of these, AtraPBP1 is highly enriched in male antennae. Fluorescence, CD and NMR studies suggest a dramatic pH-dependent conformational change, with high affinity to pheromone constituents at neutral pH and no binding at low pH.

Show MeSH
Related in: MedlinePlus