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Antagonistic peptide technology for functional dissection of CLE peptides revisited.

Czyzewicz N, Wildhagen M, Cattaneo P, Stahl Y, Pinto KG, Aalen RB, Butenko MA, Simon R, Hardtke CS, De Smet I - J. Exp. Bot. (2015)

Bottom Line: Based on the analyses, it was concluded that the antagonistic peptide approach is not the ultimate means to overcome redundancy or lack of loss-of-function lines.However, information collected using antagonistic peptide approaches (in the broad sense) can be very useful, but these approaches do not work in all cases and require a deep insight on the interaction between the ligand and its receptor to be successful.This, as well as peptide ligand structure considerations, should be taken into account before ordering a wide range of synthetic peptide variants and/or generating transgenic plants.

View Article: PubMed Central - PubMed

Affiliation: Division of Plant and Crop Sciences, School of Biosciences, University of Nottingham, Loughborough LE12 5RD, UK.

No MeSH data available.


Peptide structure. (A) Manually adjusted alignment of the 12 amino acids of CLE peptides, with IDA and IDL1 for comparison. Red stars mark peptides used in the present study, black stars those tested by Song et al. (2013). (B–D) Examples of structures of the PXXP core predicted by PEP-FOLD. (B) PGGP. The larger Thr might interfere with receptor binding, and substitution of Gly6 with Ala6 may change the angles between the Pro residues. (C) PRGP. The side chain of Arg may change direction when the Gly6 is exchanged with a Thr. (D) PSAP. A change from Ala to Thr in position six may not result in major conformational changes when Ser is present in the core. (This figure is available in colour at JXB online.)
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Figure 6: Peptide structure. (A) Manually adjusted alignment of the 12 amino acids of CLE peptides, with IDA and IDL1 for comparison. Red stars mark peptides used in the present study, black stars those tested by Song et al. (2013). (B–D) Examples of structures of the PXXP core predicted by PEP-FOLD. (B) PGGP. The larger Thr might interfere with receptor binding, and substitution of Gly6 with Ala6 may change the angles between the Pro residues. (C) PRGP. The side chain of Arg may change direction when the Gly6 is exchanged with a Thr. (D) PSAP. A change from Ala to Thr in position six may not result in major conformational changes when Ser is present in the core. (This figure is available in colour at JXB online.)

Mentions: In addition, the extent the antagonistic peptide technology can be applied to other small signalling peptides was assessed. For this, the IDA and IDA-LIKE (IDL) family were chosen, given their sequence similarity to CLEs (Stenvik et al., 2006). The IDA and IDL1 peptides of 12 amino acids share a common core at positions four to seven [PS(G/A)P] and the C-terminal end [H(N/H)] with CLV3 and some CLE peptides (Figs 5A, 6A). Like CLV3, hydroxylation of the Pro at position seven of the IDA dodecapeptide (IDAp, also referred to as PIPPo) increases the activity of the peptide (Butenko et al., 2014). An oxidative burst response in Nicotiana benthamiana can be employed as readout for the RLK HAESA-LIKE2 (HSL2) activation by exogenously applied synthetic IDA peptides (Butenko et al., 2014). Previous results indicated that IDAp binds to HSL2 with a Kd of 20nM (Butenko et al., 2014). As the wild-type IDA peptide has an Ala at position six corresponding to the Gly at that position in CLV3, and the ida mutant phenotype can be fully rescued by IDL1, which has a Gly at this position (Stenvik et al., 2008) (Fig. 6A); both of these small amino acids are evidently compatible with high signalling activity. It was, however, conceivable that substitution to the larger Thr (mIDAp6Thr) (Fig. 5A) could have an effect on receptor binding and/or activation. Therefore, the activity of mIDAp6Thr in comparison with the activity of synthetic IDAp was assessed in an oxidative burst assay. For all peptide concentrations tested, mIDAp6Thr gave the same response as IDAp in the presence of its receptor HSL2 (Fig. 5B), indicating that the mutated peptide was just as active as its wild-type counterpart. In conclusion, this mutation neither produced a ligand with weaker activity, nor a peptide with antagonistic effect.


Antagonistic peptide technology for functional dissection of CLE peptides revisited.

Czyzewicz N, Wildhagen M, Cattaneo P, Stahl Y, Pinto KG, Aalen RB, Butenko MA, Simon R, Hardtke CS, De Smet I - J. Exp. Bot. (2015)

Peptide structure. (A) Manually adjusted alignment of the 12 amino acids of CLE peptides, with IDA and IDL1 for comparison. Red stars mark peptides used in the present study, black stars those tested by Song et al. (2013). (B–D) Examples of structures of the PXXP core predicted by PEP-FOLD. (B) PGGP. The larger Thr might interfere with receptor binding, and substitution of Gly6 with Ala6 may change the angles between the Pro residues. (C) PRGP. The side chain of Arg may change direction when the Gly6 is exchanged with a Thr. (D) PSAP. A change from Ala to Thr in position six may not result in major conformational changes when Ser is present in the core. (This figure is available in colour at JXB online.)
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License 1 - License 2
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Figure 6: Peptide structure. (A) Manually adjusted alignment of the 12 amino acids of CLE peptides, with IDA and IDL1 for comparison. Red stars mark peptides used in the present study, black stars those tested by Song et al. (2013). (B–D) Examples of structures of the PXXP core predicted by PEP-FOLD. (B) PGGP. The larger Thr might interfere with receptor binding, and substitution of Gly6 with Ala6 may change the angles between the Pro residues. (C) PRGP. The side chain of Arg may change direction when the Gly6 is exchanged with a Thr. (D) PSAP. A change from Ala to Thr in position six may not result in major conformational changes when Ser is present in the core. (This figure is available in colour at JXB online.)
Mentions: In addition, the extent the antagonistic peptide technology can be applied to other small signalling peptides was assessed. For this, the IDA and IDA-LIKE (IDL) family were chosen, given their sequence similarity to CLEs (Stenvik et al., 2006). The IDA and IDL1 peptides of 12 amino acids share a common core at positions four to seven [PS(G/A)P] and the C-terminal end [H(N/H)] with CLV3 and some CLE peptides (Figs 5A, 6A). Like CLV3, hydroxylation of the Pro at position seven of the IDA dodecapeptide (IDAp, also referred to as PIPPo) increases the activity of the peptide (Butenko et al., 2014). An oxidative burst response in Nicotiana benthamiana can be employed as readout for the RLK HAESA-LIKE2 (HSL2) activation by exogenously applied synthetic IDA peptides (Butenko et al., 2014). Previous results indicated that IDAp binds to HSL2 with a Kd of 20nM (Butenko et al., 2014). As the wild-type IDA peptide has an Ala at position six corresponding to the Gly at that position in CLV3, and the ida mutant phenotype can be fully rescued by IDL1, which has a Gly at this position (Stenvik et al., 2008) (Fig. 6A); both of these small amino acids are evidently compatible with high signalling activity. It was, however, conceivable that substitution to the larger Thr (mIDAp6Thr) (Fig. 5A) could have an effect on receptor binding and/or activation. Therefore, the activity of mIDAp6Thr in comparison with the activity of synthetic IDAp was assessed in an oxidative burst assay. For all peptide concentrations tested, mIDAp6Thr gave the same response as IDAp in the presence of its receptor HSL2 (Fig. 5B), indicating that the mutated peptide was just as active as its wild-type counterpart. In conclusion, this mutation neither produced a ligand with weaker activity, nor a peptide with antagonistic effect.

Bottom Line: Based on the analyses, it was concluded that the antagonistic peptide approach is not the ultimate means to overcome redundancy or lack of loss-of-function lines.However, information collected using antagonistic peptide approaches (in the broad sense) can be very useful, but these approaches do not work in all cases and require a deep insight on the interaction between the ligand and its receptor to be successful.This, as well as peptide ligand structure considerations, should be taken into account before ordering a wide range of synthetic peptide variants and/or generating transgenic plants.

View Article: PubMed Central - PubMed

Affiliation: Division of Plant and Crop Sciences, School of Biosciences, University of Nottingham, Loughborough LE12 5RD, UK.

No MeSH data available.