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Modulation of Arabidopsis and monocot root architecture by CLAVATA3/EMBRYO SURROUNDING REGION 26 peptide.

Czyzewicz N, Shi CL, Vu LD, Van De Cotte B, Hodgman C, Butenko MA, Smet ID - J. Exp. Bot. (2015)

Bottom Line: Using chemically synthesized peptide variants, it was found that CLE26 plays an important role in regulating A. thaliana root architecture and interacts with auxin signalling.In addition, through alanine scanning and in silico structural modelling, key residues in the CLE26 peptide sequence that affect its activity are pinpointed.Finally, some interesting similarities and differences regarding the role of CLE26 in regulating monocot root architecture are presented.

View Article: PubMed Central - PubMed

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

No MeSH data available.


CLE26 and auxin response/transport. (A) CLE26 expression level as determined by qPCR in auxin-treated (6h) 5-day-old wild-type seedlings. (B–D) CLE26p-treated pDR5::GUS (B), 35S::DII:VENUS (C), and pPIN1::PIN1:GFP 5-day-old seedlings continuously grown on CLE26p (D). (E) PIN1 expression level in 7-day-old seedling roots treated with CLE26p for 24h. The bar graph indicates the mean ±SE.
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Figure 8: CLE26 and auxin response/transport. (A) CLE26 expression level as determined by qPCR in auxin-treated (6h) 5-day-old wild-type seedlings. (B–D) CLE26p-treated pDR5::GUS (B), 35S::DII:VENUS (C), and pPIN1::PIN1:GFP 5-day-old seedlings continuously grown on CLE26p (D). (E) PIN1 expression level in 7-day-old seedling roots treated with CLE26p for 24h. The bar graph indicates the mean ±SE.

Mentions: Since auxin plays a dominant role in primary root growth and lateral root initiation and development (De Smet, 2012; Lavenus et al., 2013; Tian et al., 2014), it was explored whether and potentially where CLE26p would have an influence on auxin response. For this, it was first tested if CLE26 expression was regulated by auxin. qPCR analyses showed an ~4-fold increase in CLE26 expression in wild-type seedling roots following 6h of auxin treatment (Fig. 8A). Subsequently, it was tested whether CLE26p affects the auxin response marker pDR5::GUS. At 1nM CLE26p, a concentration that significantly affected primary root growth, no obvious difference in pDR5::GUS expression was observed in the primary root tip (Fig. 8B). However, at a higher concentration (1 μM), the pDR5::GUS expression level was significantly reduced (Fig. 8B). Subsequently, the AUX/IAA protein-based auxin sensor p35S::DII:VENUS (Vernoux et al., 2011; Brunoud et al., 2012) was tested upon CLE26p treatment. While a mild increase in DII:VENUS fluorescence was observed at 1nM CLE26p, there was a dramatic increase at 1 μM (Fig. 8C). In agreement with the pDR5::GUS results (Fig. 8B), this suggested an altered auxin response in the root tip. These observations are similar to what was recently observed using 10nM CLE26p on pDR5::NLS-3xVENUS for 48h (Rodriguez-Villalon et al., 2015). Next, to determine if the effect of CLE26p on the auxin response and/or distribution in the root tip could be related to CLE26p regulation of polar auxin transport, the pPIN1::PIN1:GFP marker line was used (Benková et al., 2003). The pPIN1::PIN1:GFP marker displayed mildly and strongly reduced fluorescence at 1nM and 1 μM, respectively (Fig. 8D; Supplementary Fig. S8 at JXB online). An even stronger effect on PIN1 fluorescence and localization was observed with 1nM CLE26pP7Hyp (Supplementary Fig. S8). However, this reduction is in contrast to the qPCR results, where CLE26p treatment did not dramatically affect PIN1 expression in the root (Fig. 8E). This suggested a possible CLE26p-mediated effect on PIN1:GFP at the protein level, which could explain the reduced auxin response in the root tip.


Modulation of Arabidopsis and monocot root architecture by CLAVATA3/EMBRYO SURROUNDING REGION 26 peptide.

Czyzewicz N, Shi CL, Vu LD, Van De Cotte B, Hodgman C, Butenko MA, Smet ID - J. Exp. Bot. (2015)

CLE26 and auxin response/transport. (A) CLE26 expression level as determined by qPCR in auxin-treated (6h) 5-day-old wild-type seedlings. (B–D) CLE26p-treated pDR5::GUS (B), 35S::DII:VENUS (C), and pPIN1::PIN1:GFP 5-day-old seedlings continuously grown on CLE26p (D). (E) PIN1 expression level in 7-day-old seedling roots treated with CLE26p for 24h. The bar graph indicates the mean ±SE.
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Figure 8: CLE26 and auxin response/transport. (A) CLE26 expression level as determined by qPCR in auxin-treated (6h) 5-day-old wild-type seedlings. (B–D) CLE26p-treated pDR5::GUS (B), 35S::DII:VENUS (C), and pPIN1::PIN1:GFP 5-day-old seedlings continuously grown on CLE26p (D). (E) PIN1 expression level in 7-day-old seedling roots treated with CLE26p for 24h. The bar graph indicates the mean ±SE.
Mentions: Since auxin plays a dominant role in primary root growth and lateral root initiation and development (De Smet, 2012; Lavenus et al., 2013; Tian et al., 2014), it was explored whether and potentially where CLE26p would have an influence on auxin response. For this, it was first tested if CLE26 expression was regulated by auxin. qPCR analyses showed an ~4-fold increase in CLE26 expression in wild-type seedling roots following 6h of auxin treatment (Fig. 8A). Subsequently, it was tested whether CLE26p affects the auxin response marker pDR5::GUS. At 1nM CLE26p, a concentration that significantly affected primary root growth, no obvious difference in pDR5::GUS expression was observed in the primary root tip (Fig. 8B). However, at a higher concentration (1 μM), the pDR5::GUS expression level was significantly reduced (Fig. 8B). Subsequently, the AUX/IAA protein-based auxin sensor p35S::DII:VENUS (Vernoux et al., 2011; Brunoud et al., 2012) was tested upon CLE26p treatment. While a mild increase in DII:VENUS fluorescence was observed at 1nM CLE26p, there was a dramatic increase at 1 μM (Fig. 8C). In agreement with the pDR5::GUS results (Fig. 8B), this suggested an altered auxin response in the root tip. These observations are similar to what was recently observed using 10nM CLE26p on pDR5::NLS-3xVENUS for 48h (Rodriguez-Villalon et al., 2015). Next, to determine if the effect of CLE26p on the auxin response and/or distribution in the root tip could be related to CLE26p regulation of polar auxin transport, the pPIN1::PIN1:GFP marker line was used (Benková et al., 2003). The pPIN1::PIN1:GFP marker displayed mildly and strongly reduced fluorescence at 1nM and 1 μM, respectively (Fig. 8D; Supplementary Fig. S8 at JXB online). An even stronger effect on PIN1 fluorescence and localization was observed with 1nM CLE26pP7Hyp (Supplementary Fig. S8). However, this reduction is in contrast to the qPCR results, where CLE26p treatment did not dramatically affect PIN1 expression in the root (Fig. 8E). This suggested a possible CLE26p-mediated effect on PIN1:GFP at the protein level, which could explain the reduced auxin response in the root tip.

Bottom Line: Using chemically synthesized peptide variants, it was found that CLE26 plays an important role in regulating A. thaliana root architecture and interacts with auxin signalling.In addition, through alanine scanning and in silico structural modelling, key residues in the CLE26 peptide sequence that affect its activity are pinpointed.Finally, some interesting similarities and differences regarding the role of CLE26 in regulating monocot root architecture are presented.

View Article: PubMed Central - PubMed

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

No MeSH data available.