Limits...
The AUXIN BINDING PROTEIN 1 is required for differential auxin responses mediating root growth.

Tromas A, Braun N, Muller P, Khodus T, Paponov IA, Palme K, Ljung K, Lee JY, Benfey P, Murray JA, Scheres B, Perrot-Rechenmann C - PLoS ONE (2009)

Bottom Line: ABP1 is also implicated in the regulation of gene expression in response to auxin.Our data support that ABP1 is a key regulator for root growth and is required for auxin-mediated responses.Differential effects of ABP1 on various auxin responses support a model in which ABP1 is the major regulator for auxin action on the cell cycle and regulates auxin-mediated gene expression and cell elongation in addition to the already well known TIR1-mediated ubiquitination pathway.

View Article: PubMed Central - PubMed

Affiliation: Institut des Sciences du Végétal, CNRS UPR2355, Université Paris Sud Orsay, Gif sur Yvette, France.

ABSTRACT

Background: In plants, the phytohormone auxin is a crucial regulator sustaining growth and development. At the cellular level, auxin is interpreted differentially in a tissue- and dose-dependent manner. Mechanisms of auxin signalling are partially unknown and the contribution of the AUXIN BINDING PROTEIN 1 (ABP1) as an auxin receptor is still a matter of debate.

Methodology/principal findings: Here we took advantage of the present knowledge of the root biological system to demonstrate that ABP1 is required for auxin response. The use of conditional ABP1 defective plants reveals that the protein is essential for maintenance of the root meristem and acts at least on the D-type CYCLIN/RETINOBLASTOMA pathway to control entry into the cell cycle. ABP1 affects PLETHORA gradients and confers auxin sensitivity to root cells thus defining the competence of the cells to be maintained within the meristem or to elongate. ABP1 is also implicated in the regulation of gene expression in response to auxin.

Conclusions/significance: Our data support that ABP1 is a key regulator for root growth and is required for auxin-mediated responses. Differential effects of ABP1 on various auxin responses support a model in which ABP1 is the major regulator for auxin action on the cell cycle and regulates auxin-mediated gene expression and cell elongation in addition to the already well known TIR1-mediated ubiquitination pathway.

Show MeSH

Related in: MedlinePlus

Inactivation of ABP1 leads to consumption of meristematic cells.A–F, Histology of root apex visualized on optical longitudinal sections of living roots stained with FM4-64 (in red). A,B, Col0; C,D, SS12K; E,F, ABP1-AS of 4 day old seedlings grown in the absence (A,C,E) or in the presence of ethanol vapour (B,D,F). Scale bar 40 µm. G–J, Morphometric analysis comparing ethanol induced (blue bars) and non induced (grey bars) ABP1AS at 3dpg. G, meristematic zone length measured from the QC to the end of the lateral root cap; H, cortex cell number in the meristematic zone; I, root tip diameter measured at the end of the lateral root cap; J, epidermal and cortical cell length of differentiated cells, measured after the emergence of root hairs. Standard deviation were calculated from sample number >40. K–L, Optical radial sections of non induced (K) and ethanol induced (L) SS12K roots taken below the end of the lateral root cap. M–N, Close up of the root tip organisation with a focus on the QC and the columella of SS12K not induced (M) or ethanol induced (N). Scale bar 40 µm. O–P, S17:GFP marker AT2G22850 of differentiated phloem pole pericycle (in green) in non induced (O) and induced (P) SS12K seedlings at 4 dpg. Scale bar 80 µm. Q–V, Patterning of cell type specific markers in induced control (Q, S, U) and ABP1-AS (R, T, V) roots. Protophloem S32:GFP AT2G18380 (Q, R); protoxylem S4:GFP AT3G25710 (S, T) and root cap and procambium A8:GFP AT3G48100 (U, V) [16]. Scale bar 40 µm. GFP marker lines [16] were introgressed into SS12K and ABP1AS plants and double homozygous plants were selected from F3 progeny. No significant changes of GFP (in green) expression pattern were detected between ethanol induced or non induced ABP1AS plants or in comparison with reported expression in wild-type background.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2744284&req=5

pone-0006648-g002: Inactivation of ABP1 leads to consumption of meristematic cells.A–F, Histology of root apex visualized on optical longitudinal sections of living roots stained with FM4-64 (in red). A,B, Col0; C,D, SS12K; E,F, ABP1-AS of 4 day old seedlings grown in the absence (A,C,E) or in the presence of ethanol vapour (B,D,F). Scale bar 40 µm. G–J, Morphometric analysis comparing ethanol induced (blue bars) and non induced (grey bars) ABP1AS at 3dpg. G, meristematic zone length measured from the QC to the end of the lateral root cap; H, cortex cell number in the meristematic zone; I, root tip diameter measured at the end of the lateral root cap; J, epidermal and cortical cell length of differentiated cells, measured after the emergence of root hairs. Standard deviation were calculated from sample number >40. K–L, Optical radial sections of non induced (K) and ethanol induced (L) SS12K roots taken below the end of the lateral root cap. M–N, Close up of the root tip organisation with a focus on the QC and the columella of SS12K not induced (M) or ethanol induced (N). Scale bar 40 µm. O–P, S17:GFP marker AT2G22850 of differentiated phloem pole pericycle (in green) in non induced (O) and induced (P) SS12K seedlings at 4 dpg. Scale bar 80 µm. Q–V, Patterning of cell type specific markers in induced control (Q, S, U) and ABP1-AS (R, T, V) roots. Protophloem S32:GFP AT2G18380 (Q, R); protoxylem S4:GFP AT3G25710 (S, T) and root cap and procambium A8:GFP AT3G48100 (U, V) [16]. Scale bar 40 µm. GFP marker lines [16] were introgressed into SS12K and ABP1AS plants and double homozygous plants were selected from F3 progeny. No significant changes of GFP (in green) expression pattern were detected between ethanol induced or non induced ABP1AS plants or in comparison with reported expression in wild-type background.

Mentions: To circumvent the embryo-lethality of ABP1 knock-out [7], we used ethanol inducible conditional knock-down plants generated via an antisense ABP1 construct to decrease its expression (ABP1AS lines) or via cellular immunization to inactivate ABP1 protein through its in vivo interaction with the recombinant antibody scFv12. The latter recognizes a conformational epitope of ABP1 overlapping the auxin binding site (SS12S and SS12K lines) thus impairing the capacity of the protein to bind and respond to auxin [10], [15]. The recombinant antibody was detected in enriched microsomal samples of ethanol induced SS12K (Figure 1A) and we showed by reciprocal co-immunoprecipitation experiments that the scFv12 produced in Arabidopsis interacts with AtABP1 in vivo (Figure 1B). ABP1 was still detected in root samples expressing the scFv12 whereas the protein was not detected in induced antisense samples (Figure 1C). At three days post germination (dpg), ethanol induced SS12S, SS12K and ABP1AS plants exhibited similar phenotypes displaying drastic root growth reduction of 60 to 80% compared to ethanol induced control plants (Figure 1D–H). To determine which cellular alterations were responsible for such severe root growth defect, we performed a detailed analysis of SS12K and ABP1AS primary roots. The size of the meristem of ABP1 inactivated roots is about one third of that of controls which correlates with a reduced number of meristematic cells (Figures 2 A–H). Differentiated cortical cells reach a similar length as in control roots (Figure 2J), indicating that longitudinal elongation is not defective in ABP1 inactivated plants. The root diameter is, however, reduced by more than 40% due to decreased radial expansion but the radial tissue organisation inherited from the embryonic root pattern is unaltered (Figure 2I, K, L). Introgression of a series of specific cell type GFP marker lines [16] confirmed maintenance of radial patterns (Figure 2Q–V). At the root apex, a cell layer is missing in both columella and lateral root cap in more then 80% of roots with repressed ABP1 activity (Figure 2M–N). Changes in the longitudinal gradient of root differentiation was confirmed by the use of the S17 GFP marker [16], which is expressed in phloem pole pericycle cells and is detected in the differentiation zone (Figure 2O). After ABP1 inactivation, S17 marker expression is observed at a more distal position, indicating that cells that have left the meristem rapidly begin differentiation (Figure 2P).


The AUXIN BINDING PROTEIN 1 is required for differential auxin responses mediating root growth.

Tromas A, Braun N, Muller P, Khodus T, Paponov IA, Palme K, Ljung K, Lee JY, Benfey P, Murray JA, Scheres B, Perrot-Rechenmann C - PLoS ONE (2009)

Inactivation of ABP1 leads to consumption of meristematic cells.A–F, Histology of root apex visualized on optical longitudinal sections of living roots stained with FM4-64 (in red). A,B, Col0; C,D, SS12K; E,F, ABP1-AS of 4 day old seedlings grown in the absence (A,C,E) or in the presence of ethanol vapour (B,D,F). Scale bar 40 µm. G–J, Morphometric analysis comparing ethanol induced (blue bars) and non induced (grey bars) ABP1AS at 3dpg. G, meristematic zone length measured from the QC to the end of the lateral root cap; H, cortex cell number in the meristematic zone; I, root tip diameter measured at the end of the lateral root cap; J, epidermal and cortical cell length of differentiated cells, measured after the emergence of root hairs. Standard deviation were calculated from sample number >40. K–L, Optical radial sections of non induced (K) and ethanol induced (L) SS12K roots taken below the end of the lateral root cap. M–N, Close up of the root tip organisation with a focus on the QC and the columella of SS12K not induced (M) or ethanol induced (N). Scale bar 40 µm. O–P, S17:GFP marker AT2G22850 of differentiated phloem pole pericycle (in green) in non induced (O) and induced (P) SS12K seedlings at 4 dpg. Scale bar 80 µm. Q–V, Patterning of cell type specific markers in induced control (Q, S, U) and ABP1-AS (R, T, V) roots. Protophloem S32:GFP AT2G18380 (Q, R); protoxylem S4:GFP AT3G25710 (S, T) and root cap and procambium A8:GFP AT3G48100 (U, V) [16]. Scale bar 40 µm. GFP marker lines [16] were introgressed into SS12K and ABP1AS plants and double homozygous plants were selected from F3 progeny. No significant changes of GFP (in green) expression pattern were detected between ethanol induced or non induced ABP1AS plants or in comparison with reported expression in wild-type background.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2744284&req=5

pone-0006648-g002: Inactivation of ABP1 leads to consumption of meristematic cells.A–F, Histology of root apex visualized on optical longitudinal sections of living roots stained with FM4-64 (in red). A,B, Col0; C,D, SS12K; E,F, ABP1-AS of 4 day old seedlings grown in the absence (A,C,E) or in the presence of ethanol vapour (B,D,F). Scale bar 40 µm. G–J, Morphometric analysis comparing ethanol induced (blue bars) and non induced (grey bars) ABP1AS at 3dpg. G, meristematic zone length measured from the QC to the end of the lateral root cap; H, cortex cell number in the meristematic zone; I, root tip diameter measured at the end of the lateral root cap; J, epidermal and cortical cell length of differentiated cells, measured after the emergence of root hairs. Standard deviation were calculated from sample number >40. K–L, Optical radial sections of non induced (K) and ethanol induced (L) SS12K roots taken below the end of the lateral root cap. M–N, Close up of the root tip organisation with a focus on the QC and the columella of SS12K not induced (M) or ethanol induced (N). Scale bar 40 µm. O–P, S17:GFP marker AT2G22850 of differentiated phloem pole pericycle (in green) in non induced (O) and induced (P) SS12K seedlings at 4 dpg. Scale bar 80 µm. Q–V, Patterning of cell type specific markers in induced control (Q, S, U) and ABP1-AS (R, T, V) roots. Protophloem S32:GFP AT2G18380 (Q, R); protoxylem S4:GFP AT3G25710 (S, T) and root cap and procambium A8:GFP AT3G48100 (U, V) [16]. Scale bar 40 µm. GFP marker lines [16] were introgressed into SS12K and ABP1AS plants and double homozygous plants were selected from F3 progeny. No significant changes of GFP (in green) expression pattern were detected between ethanol induced or non induced ABP1AS plants or in comparison with reported expression in wild-type background.
Mentions: To circumvent the embryo-lethality of ABP1 knock-out [7], we used ethanol inducible conditional knock-down plants generated via an antisense ABP1 construct to decrease its expression (ABP1AS lines) or via cellular immunization to inactivate ABP1 protein through its in vivo interaction with the recombinant antibody scFv12. The latter recognizes a conformational epitope of ABP1 overlapping the auxin binding site (SS12S and SS12K lines) thus impairing the capacity of the protein to bind and respond to auxin [10], [15]. The recombinant antibody was detected in enriched microsomal samples of ethanol induced SS12K (Figure 1A) and we showed by reciprocal co-immunoprecipitation experiments that the scFv12 produced in Arabidopsis interacts with AtABP1 in vivo (Figure 1B). ABP1 was still detected in root samples expressing the scFv12 whereas the protein was not detected in induced antisense samples (Figure 1C). At three days post germination (dpg), ethanol induced SS12S, SS12K and ABP1AS plants exhibited similar phenotypes displaying drastic root growth reduction of 60 to 80% compared to ethanol induced control plants (Figure 1D–H). To determine which cellular alterations were responsible for such severe root growth defect, we performed a detailed analysis of SS12K and ABP1AS primary roots. The size of the meristem of ABP1 inactivated roots is about one third of that of controls which correlates with a reduced number of meristematic cells (Figures 2 A–H). Differentiated cortical cells reach a similar length as in control roots (Figure 2J), indicating that longitudinal elongation is not defective in ABP1 inactivated plants. The root diameter is, however, reduced by more than 40% due to decreased radial expansion but the radial tissue organisation inherited from the embryonic root pattern is unaltered (Figure 2I, K, L). Introgression of a series of specific cell type GFP marker lines [16] confirmed maintenance of radial patterns (Figure 2Q–V). At the root apex, a cell layer is missing in both columella and lateral root cap in more then 80% of roots with repressed ABP1 activity (Figure 2M–N). Changes in the longitudinal gradient of root differentiation was confirmed by the use of the S17 GFP marker [16], which is expressed in phloem pole pericycle cells and is detected in the differentiation zone (Figure 2O). After ABP1 inactivation, S17 marker expression is observed at a more distal position, indicating that cells that have left the meristem rapidly begin differentiation (Figure 2P).

Bottom Line: ABP1 is also implicated in the regulation of gene expression in response to auxin.Our data support that ABP1 is a key regulator for root growth and is required for auxin-mediated responses.Differential effects of ABP1 on various auxin responses support a model in which ABP1 is the major regulator for auxin action on the cell cycle and regulates auxin-mediated gene expression and cell elongation in addition to the already well known TIR1-mediated ubiquitination pathway.

View Article: PubMed Central - PubMed

Affiliation: Institut des Sciences du Végétal, CNRS UPR2355, Université Paris Sud Orsay, Gif sur Yvette, France.

ABSTRACT

Background: In plants, the phytohormone auxin is a crucial regulator sustaining growth and development. At the cellular level, auxin is interpreted differentially in a tissue- and dose-dependent manner. Mechanisms of auxin signalling are partially unknown and the contribution of the AUXIN BINDING PROTEIN 1 (ABP1) as an auxin receptor is still a matter of debate.

Methodology/principal findings: Here we took advantage of the present knowledge of the root biological system to demonstrate that ABP1 is required for auxin response. The use of conditional ABP1 defective plants reveals that the protein is essential for maintenance of the root meristem and acts at least on the D-type CYCLIN/RETINOBLASTOMA pathway to control entry into the cell cycle. ABP1 affects PLETHORA gradients and confers auxin sensitivity to root cells thus defining the competence of the cells to be maintained within the meristem or to elongate. ABP1 is also implicated in the regulation of gene expression in response to auxin.

Conclusions/significance: Our data support that ABP1 is a key regulator for root growth and is required for auxin-mediated responses. Differential effects of ABP1 on various auxin responses support a model in which ABP1 is the major regulator for auxin action on the cell cycle and regulates auxin-mediated gene expression and cell elongation in addition to the already well known TIR1-mediated ubiquitination pathway.

Show MeSH
Related in: MedlinePlus