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Inhibition of cell expansion by rapid ABP1-mediated auxin effect on microtubules.

Chen X, Grandont L, Li H, Hauschild R, Paque S, Abuzeineh A, Rakusová H, Benkova E, Perrot-Rechenmann C, Friml J - Nature (2014)

Bottom Line: Here we show in the model plant Arabidopsis thaliana that in elongating cells exogenous application of auxin or redistribution of endogenous auxin induces very rapid microtubule re-orientation from transverse to longitudinal, coherent with the inhibition of cell expansion.This fast auxin effect requires auxin binding protein 1 (ABP1) and involves a contribution of downstream signalling components such as ROP6 GTPase, ROP-interactive protein RIC1 and the microtubule-severing protein katanin.These components are required for rapid auxin- and ABP1-mediated re-orientation of microtubules to regulate cell elongation in roots and dark-grown hypocotyls as well as asymmetric growth during gravitropic responses.

View Article: PubMed Central - PubMed

Affiliation: 1] Institute of Science and Technology Austria (IST Austria), Am Campus 1, 3400 Klosterneuburg, Austria [2] Department of Plant Systems Biology, Vlaams Instituut voor Biotechnologie (VIB), Ghent University, B-9052 Gent, Belgium [3] Department of Plant Biotechnology and Genetics, Ghent University, B-9052 Gent, Belgium.

ABSTRACT
The prominent and evolutionarily ancient role of the plant hormone auxin is the regulation of cell expansion. Cell expansion requires ordered arrangement of the cytoskeleton but molecular mechanisms underlying its regulation by signalling molecules including auxin are unknown. Here we show in the model plant Arabidopsis thaliana that in elongating cells exogenous application of auxin or redistribution of endogenous auxin induces very rapid microtubule re-orientation from transverse to longitudinal, coherent with the inhibition of cell expansion. This fast auxin effect requires auxin binding protein 1 (ABP1) and involves a contribution of downstream signalling components such as ROP6 GTPase, ROP-interactive protein RIC1 and the microtubule-severing protein katanin. These components are required for rapid auxin- and ABP1-mediated re-orientation of microtubules to regulate cell elongation in roots and dark-grown hypocotyls as well as asymmetric growth during gravitropic responses.

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Auxin-ABP1 controls MT arrangement through the downstream ROP6-RIC1-KTN1 signaling(a) MAP4-GFP visualization of MTs orientation in the root of WT, rop6-1, ric1-1, SS12S ric1-1, SS12K ric1-1 following DMSO application for 60 min. Corresponding to quantifications in Fig. 4a.(b-c) MTs reorientation patterns were visualized by MAP4-GFP in the roots of WT and rop6-1+/− following DMSO or 100nM NAA application for 60 min (Student’s T-test, p>0.05).(d) The transcript level of the scFv12 coding the recombinant antibody responsible for ABP1 knockdown in WT, ric1-1, ktn1, SS12S, SS12K, SS12S ric1-1, SS12K ric1-1, SS12S ktn1 and SS12K ktn1 after 48h ethanol induction. The transcript level of the scFv12 in SS12S was standardized as “1” (Student’s T-test, p>0.05).(e) MTs orientation by MAP4-GFP in dark grown hypocotyls of WT, SS12K, ktn1, SS12K ktn1 (with 24h ethanol induction) following DMSO application for 60 min. Corresponding to Fig. 4b. In all panels, error bars are s.e.m. Scale bars: 5μm (a-b) and 10μm (d).
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Figure 11: Auxin-ABP1 controls MT arrangement through the downstream ROP6-RIC1-KTN1 signaling(a) MAP4-GFP visualization of MTs orientation in the root of WT, rop6-1, ric1-1, SS12S ric1-1, SS12K ric1-1 following DMSO application for 60 min. Corresponding to quantifications in Fig. 4a.(b-c) MTs reorientation patterns were visualized by MAP4-GFP in the roots of WT and rop6-1+/− following DMSO or 100nM NAA application for 60 min (Student’s T-test, p>0.05).(d) The transcript level of the scFv12 coding the recombinant antibody responsible for ABP1 knockdown in WT, ric1-1, ktn1, SS12S, SS12K, SS12S ric1-1, SS12K ric1-1, SS12S ktn1 and SS12K ktn1 after 48h ethanol induction. The transcript level of the scFv12 in SS12S was standardized as “1” (Student’s T-test, p>0.05).(e) MTs orientation by MAP4-GFP in dark grown hypocotyls of WT, SS12K, ktn1, SS12K ktn1 (with 24h ethanol induction) following DMSO application for 60 min. Corresponding to Fig. 4b. In all panels, error bars are s.e.m. Scale bars: 5μm (a-b) and 10μm (d).

Mentions: Next we addressed the downstream mechanism by which ABP1 mediates the auxin effect on MT arrangement. Although auxin induces calcium transients22, the manipulation of exogenous calcium had very different effects on MT arrangements as compared to auxin (Extended Data Fig. 6). Next we tested the downstream components of ABP1 pathway: the ROP6 GTPase, its effector RIC116 and its downstream component MT severing protein KTN123,24. We analyzed MTs in rop6-1, ric1-1 and ktn1 mutants. Compared to WT, roots of rop6-1 and ric1-1 showed almost normal transversal MTs but were much less auxin responsive (Fig. 4a, Extended Data Fig. 7a). Double mutants (SS12S ric1-1, SS12K ric1-1), with ABP1 inactivation, exhibited SS12K/S root phenotype15 but ric1-1 MT arrangement (Fig. 4a, Extended Data Fig. 7a-d), consistent with RIC1 reported action downstream of ABP1 in early responses25. ktn1 mutant exhibited severe MTs phenotype with completely random MTs in root compromising further analysis. MTs were less disturbed in dark grown ktn1 hypocotyl allowing investigating the response to auxin and genetic interaction with ABP1 (Extended Data Fig. 7e). Rapid auxin-induced reorientation of MTs was impaired in ktn1 (Fig. 4b). Inactivation of ABP1 in ktn1 (SS12K ktn1) resulted in a MT pattern similar to SS12K and confers insensitivity to auxin (Fig. 4b). These data suggest that KTN1 is required for MTs reorientation in response to auxin but that other MT associated components might be involved as well. The present data and the crucial roles of RIC1 and KTN1 in the control of microtubule architecture and crossover6,24 support that auxin-dependent ABP1 signaling might act through Rho GTPases and RIC effectors on critical targets as KTN1 for guiding MT orientation.


Inhibition of cell expansion by rapid ABP1-mediated auxin effect on microtubules.

Chen X, Grandont L, Li H, Hauschild R, Paque S, Abuzeineh A, Rakusová H, Benkova E, Perrot-Rechenmann C, Friml J - Nature (2014)

Auxin-ABP1 controls MT arrangement through the downstream ROP6-RIC1-KTN1 signaling(a) MAP4-GFP visualization of MTs orientation in the root of WT, rop6-1, ric1-1, SS12S ric1-1, SS12K ric1-1 following DMSO application for 60 min. Corresponding to quantifications in Fig. 4a.(b-c) MTs reorientation patterns were visualized by MAP4-GFP in the roots of WT and rop6-1+/− following DMSO or 100nM NAA application for 60 min (Student’s T-test, p>0.05).(d) The transcript level of the scFv12 coding the recombinant antibody responsible for ABP1 knockdown in WT, ric1-1, ktn1, SS12S, SS12K, SS12S ric1-1, SS12K ric1-1, SS12S ktn1 and SS12K ktn1 after 48h ethanol induction. The transcript level of the scFv12 in SS12S was standardized as “1” (Student’s T-test, p>0.05).(e) MTs orientation by MAP4-GFP in dark grown hypocotyls of WT, SS12K, ktn1, SS12K ktn1 (with 24h ethanol induction) following DMSO application for 60 min. Corresponding to Fig. 4b. In all panels, error bars are s.e.m. Scale bars: 5μm (a-b) and 10μm (d).
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Figure 11: Auxin-ABP1 controls MT arrangement through the downstream ROP6-RIC1-KTN1 signaling(a) MAP4-GFP visualization of MTs orientation in the root of WT, rop6-1, ric1-1, SS12S ric1-1, SS12K ric1-1 following DMSO application for 60 min. Corresponding to quantifications in Fig. 4a.(b-c) MTs reorientation patterns were visualized by MAP4-GFP in the roots of WT and rop6-1+/− following DMSO or 100nM NAA application for 60 min (Student’s T-test, p>0.05).(d) The transcript level of the scFv12 coding the recombinant antibody responsible for ABP1 knockdown in WT, ric1-1, ktn1, SS12S, SS12K, SS12S ric1-1, SS12K ric1-1, SS12S ktn1 and SS12K ktn1 after 48h ethanol induction. The transcript level of the scFv12 in SS12S was standardized as “1” (Student’s T-test, p>0.05).(e) MTs orientation by MAP4-GFP in dark grown hypocotyls of WT, SS12K, ktn1, SS12K ktn1 (with 24h ethanol induction) following DMSO application for 60 min. Corresponding to Fig. 4b. In all panels, error bars are s.e.m. Scale bars: 5μm (a-b) and 10μm (d).
Mentions: Next we addressed the downstream mechanism by which ABP1 mediates the auxin effect on MT arrangement. Although auxin induces calcium transients22, the manipulation of exogenous calcium had very different effects on MT arrangements as compared to auxin (Extended Data Fig. 6). Next we tested the downstream components of ABP1 pathway: the ROP6 GTPase, its effector RIC116 and its downstream component MT severing protein KTN123,24. We analyzed MTs in rop6-1, ric1-1 and ktn1 mutants. Compared to WT, roots of rop6-1 and ric1-1 showed almost normal transversal MTs but were much less auxin responsive (Fig. 4a, Extended Data Fig. 7a). Double mutants (SS12S ric1-1, SS12K ric1-1), with ABP1 inactivation, exhibited SS12K/S root phenotype15 but ric1-1 MT arrangement (Fig. 4a, Extended Data Fig. 7a-d), consistent with RIC1 reported action downstream of ABP1 in early responses25. ktn1 mutant exhibited severe MTs phenotype with completely random MTs in root compromising further analysis. MTs were less disturbed in dark grown ktn1 hypocotyl allowing investigating the response to auxin and genetic interaction with ABP1 (Extended Data Fig. 7e). Rapid auxin-induced reorientation of MTs was impaired in ktn1 (Fig. 4b). Inactivation of ABP1 in ktn1 (SS12K ktn1) resulted in a MT pattern similar to SS12K and confers insensitivity to auxin (Fig. 4b). These data suggest that KTN1 is required for MTs reorientation in response to auxin but that other MT associated components might be involved as well. The present data and the crucial roles of RIC1 and KTN1 in the control of microtubule architecture and crossover6,24 support that auxin-dependent ABP1 signaling might act through Rho GTPases and RIC effectors on critical targets as KTN1 for guiding MT orientation.

Bottom Line: Here we show in the model plant Arabidopsis thaliana that in elongating cells exogenous application of auxin or redistribution of endogenous auxin induces very rapid microtubule re-orientation from transverse to longitudinal, coherent with the inhibition of cell expansion.This fast auxin effect requires auxin binding protein 1 (ABP1) and involves a contribution of downstream signalling components such as ROP6 GTPase, ROP-interactive protein RIC1 and the microtubule-severing protein katanin.These components are required for rapid auxin- and ABP1-mediated re-orientation of microtubules to regulate cell elongation in roots and dark-grown hypocotyls as well as asymmetric growth during gravitropic responses.

View Article: PubMed Central - PubMed

Affiliation: 1] Institute of Science and Technology Austria (IST Austria), Am Campus 1, 3400 Klosterneuburg, Austria [2] Department of Plant Systems Biology, Vlaams Instituut voor Biotechnologie (VIB), Ghent University, B-9052 Gent, Belgium [3] Department of Plant Biotechnology and Genetics, Ghent University, B-9052 Gent, Belgium.

ABSTRACT
The prominent and evolutionarily ancient role of the plant hormone auxin is the regulation of cell expansion. Cell expansion requires ordered arrangement of the cytoskeleton but molecular mechanisms underlying its regulation by signalling molecules including auxin are unknown. Here we show in the model plant Arabidopsis thaliana that in elongating cells exogenous application of auxin or redistribution of endogenous auxin induces very rapid microtubule re-orientation from transverse to longitudinal, coherent with the inhibition of cell expansion. This fast auxin effect requires auxin binding protein 1 (ABP1) and involves a contribution of downstream signalling components such as ROP6 GTPase, ROP-interactive protein RIC1 and the microtubule-severing protein katanin. These components are required for rapid auxin- and ABP1-mediated re-orientation of microtubules to regulate cell elongation in roots and dark-grown hypocotyls as well as asymmetric growth during gravitropic responses.

Show MeSH