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Chemosensillum immunolocalization and ligand specificity of chemosensory proteins in the alfalfa plant bug Adelphocoris lineolatus (Goeze).

Sun L, Zhou JJ, Gu SH, Xiao HJ, Guo YY, Liu ZW, Zhang YJ - Sci Rep (2015)

Bottom Line: Further cellular investigation of their immunolocalization revealed their dynamic protein expression profiles among different types of antennal sensilla.In ad`dition, a cooperative interaction was observed between two co-expressed CSPs resulting in an increase of the binding affinities by a mixture of AlinCSP5 and AlinCSP6 to terpenoids which do not bind to individual CSPs.These findings in combination with our previous data for AlinCSP1-3 indicate a possible functional differentiation of CSPs in the A. lineolatus olfactory system.

View Article: PubMed Central - PubMed

Affiliation: 1] State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing, 100193, China [2] Key Laboratory of Tea Plants Biology and Resources Utilization of Agriculture Ministry, Tea Research Institute, Chinese Academy of Agricultural Sciences, Hangzhou, 310008, China [3] Key Laboratory of Integrated Management of Crop Diseases and Pests (Ministry of Education), College of Plant Protection, Nanjing Agricultural University, Nanjing, 210095, China.

ABSTRACT
Insect chemosensory proteins (CSPs) are a family of small soluble proteins. To date, their physiological functions in insect olfaction remain largely controversial in comparison to odorant binding proteins (OBPs). In present study, we reported the antenna specific expression of three CSPs (AlinCSP4-6) from Adelphocoris lineolatus, their distinct chemosensillum distribution as well as ligand binding capability thus providing the evidence for the possible roles that they could play in semiochemical detection of the plant bug A. lineolatus. The results of qRT-PCR and western blot assay clearly showed that all of these three CSPs are highly expressed in the adult antennae, the olfactory organ of insects. Further cellular investigation of their immunolocalization revealed their dynamic protein expression profiles among different types of antennal sensilla. In a fluorescence competitive binding assay, the selective ligand binding was observed for AlinCSP4-6. In ad`dition, a cooperative interaction was observed between two co-expressed CSPs resulting in an increase of the binding affinities by a mixture of AlinCSP5 and AlinCSP6 to terpenoids which do not bind to individual CSPs. These findings in combination with our previous data for AlinCSP1-3 indicate a possible functional differentiation of CSPs in the A. lineolatus olfactory system.

No MeSH data available.


Relative transcript levels of AlinCSP4-6 among different adult tissues of both sexes analyzed by qPCR.The fold changes are relative to the transcript levels in the abdomen. The error bars represents the standard errors, and the different letters (a, b and α, β) indicate significant differences (p < 0.05) among different tissues in male and female, respectively.
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f2: Relative transcript levels of AlinCSP4-6 among different adult tissues of both sexes analyzed by qPCR.The fold changes are relative to the transcript levels in the abdomen. The error bars represents the standard errors, and the different letters (a, b and α, β) indicate significant differences (p < 0.05) among different tissues in male and female, respectively.

Mentions: Three full-length cDNA sequences that encode for putative chemosensory proteins were identified with Blastx and CSP motif search against the antenna cDNA library of A. lineolatus, named as AlinCSP4, AlinCSP5, and AlinCSP6 and deposited in GenBank with the accession numbers of GQ477017, GQ477018, GQ477019, respectively. AlinCSP4-6 genes contain open reading frames (ORF) of 384 bp, 384 bp, and 339 bp, respectively. The predicted amino acid sequences of AlinCSP4-6 share the conserved characteristics of typical insect CSP family, such as the typical four highly conserved cysteines and spacing between them, and the predicted signal peptides of 19, 21, and 19 amino acids at the N-terminus, respectively (Figure S1). The calculated molecular masses of mature AlinCSP4-6 proteins are 12.59 kDa, 12.59 kDa, and 13.29 kDa, respectively. The predicted isoelectric points of the mature AlinCSP4-6 proteins are 4.92, 6.92, and 5.70, respectively. The amino acid identity among AlinCSP4-6 is approximately 41% overall, and their sequence alignment with 31 CSP sequences from other insect species indicates a very divergent signal peptide region and some highly conserved amino acid residues in addition to the four conserved cysteine residues (Figure 1). It is well known that olfactory genes would be expressed highly in the olfactory tissue namely antennae. Therefore, the quantitative real time RT-PCR (qRT-PCR) was conducted to investigate the tissue distribution and relative expression levels of AlinCSP4-6 transcripts in both male and female antennae. The results revealed that all of these three AlinCSP genes are significantly but not exclusively expressed in the adult antennae in both sexes (Figure 2).


Chemosensillum immunolocalization and ligand specificity of chemosensory proteins in the alfalfa plant bug Adelphocoris lineolatus (Goeze).

Sun L, Zhou JJ, Gu SH, Xiao HJ, Guo YY, Liu ZW, Zhang YJ - Sci Rep (2015)

Relative transcript levels of AlinCSP4-6 among different adult tissues of both sexes analyzed by qPCR.The fold changes are relative to the transcript levels in the abdomen. The error bars represents the standard errors, and the different letters (a, b and α, β) indicate significant differences (p < 0.05) among different tissues in male and female, respectively.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4308698&req=5

f2: Relative transcript levels of AlinCSP4-6 among different adult tissues of both sexes analyzed by qPCR.The fold changes are relative to the transcript levels in the abdomen. The error bars represents the standard errors, and the different letters (a, b and α, β) indicate significant differences (p < 0.05) among different tissues in male and female, respectively.
Mentions: Three full-length cDNA sequences that encode for putative chemosensory proteins were identified with Blastx and CSP motif search against the antenna cDNA library of A. lineolatus, named as AlinCSP4, AlinCSP5, and AlinCSP6 and deposited in GenBank with the accession numbers of GQ477017, GQ477018, GQ477019, respectively. AlinCSP4-6 genes contain open reading frames (ORF) of 384 bp, 384 bp, and 339 bp, respectively. The predicted amino acid sequences of AlinCSP4-6 share the conserved characteristics of typical insect CSP family, such as the typical four highly conserved cysteines and spacing between them, and the predicted signal peptides of 19, 21, and 19 amino acids at the N-terminus, respectively (Figure S1). The calculated molecular masses of mature AlinCSP4-6 proteins are 12.59 kDa, 12.59 kDa, and 13.29 kDa, respectively. The predicted isoelectric points of the mature AlinCSP4-6 proteins are 4.92, 6.92, and 5.70, respectively. The amino acid identity among AlinCSP4-6 is approximately 41% overall, and their sequence alignment with 31 CSP sequences from other insect species indicates a very divergent signal peptide region and some highly conserved amino acid residues in addition to the four conserved cysteine residues (Figure 1). It is well known that olfactory genes would be expressed highly in the olfactory tissue namely antennae. Therefore, the quantitative real time RT-PCR (qRT-PCR) was conducted to investigate the tissue distribution and relative expression levels of AlinCSP4-6 transcripts in both male and female antennae. The results revealed that all of these three AlinCSP genes are significantly but not exclusively expressed in the adult antennae in both sexes (Figure 2).

Bottom Line: Further cellular investigation of their immunolocalization revealed their dynamic protein expression profiles among different types of antennal sensilla.In ad`dition, a cooperative interaction was observed between two co-expressed CSPs resulting in an increase of the binding affinities by a mixture of AlinCSP5 and AlinCSP6 to terpenoids which do not bind to individual CSPs.These findings in combination with our previous data for AlinCSP1-3 indicate a possible functional differentiation of CSPs in the A. lineolatus olfactory system.

View Article: PubMed Central - PubMed

Affiliation: 1] State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing, 100193, China [2] Key Laboratory of Tea Plants Biology and Resources Utilization of Agriculture Ministry, Tea Research Institute, Chinese Academy of Agricultural Sciences, Hangzhou, 310008, China [3] Key Laboratory of Integrated Management of Crop Diseases and Pests (Ministry of Education), College of Plant Protection, Nanjing Agricultural University, Nanjing, 210095, China.

ABSTRACT
Insect chemosensory proteins (CSPs) are a family of small soluble proteins. To date, their physiological functions in insect olfaction remain largely controversial in comparison to odorant binding proteins (OBPs). In present study, we reported the antenna specific expression of three CSPs (AlinCSP4-6) from Adelphocoris lineolatus, their distinct chemosensillum distribution as well as ligand binding capability thus providing the evidence for the possible roles that they could play in semiochemical detection of the plant bug A. lineolatus. The results of qRT-PCR and western blot assay clearly showed that all of these three CSPs are highly expressed in the adult antennae, the olfactory organ of insects. Further cellular investigation of their immunolocalization revealed their dynamic protein expression profiles among different types of antennal sensilla. In a fluorescence competitive binding assay, the selective ligand binding was observed for AlinCSP4-6. In ad`dition, a cooperative interaction was observed between two co-expressed CSPs resulting in an increase of the binding affinities by a mixture of AlinCSP5 and AlinCSP6 to terpenoids which do not bind to individual CSPs. These findings in combination with our previous data for AlinCSP1-3 indicate a possible functional differentiation of CSPs in the A. lineolatus olfactory system.

No MeSH data available.