Extracellular adenosine triphosphate (eATP) is an apoplastic signaling molecule that plays an essential role in the growth and development of plants. Arabidopsis seedlings have been reported to respond to eATP; however, the downstream signaling components are still not well understood. In this study, we report that an ethylene-responsive factor, Redox-Responsive Transcription Factor 1 (RRTF1), is involved in eATP-regulated Arabidopsis thaliana seedling growth. Exogenous adenosine triphosphate inhibited green seedling root growth and induced hypocotyl bending of etiolated seedlings. RRTF1 loss-of-function mutant (rrtf1) seedlings showed decreased responses to eATP, while its complementation or overexpression led to recovered or increased eATP responsiveness. RRTF1 was expressed rapidly after eATP stimulation and then migrated into the nuclei of root tip cells. eATP-induced auxin accumulation in root tip or hypocotyl cells was impaired in rrtf1. Chromatin immunoprecipitation and high-throughput sequencing results indicated that eATP induced some genes related to cell growth and development in wild type but not in rrtf1 cells. These results suggest that RRTF1 may be involved in eATP signaling by regulating functional gene expression and cell metabolism in Arabidopsis seedlings.
Extracellular ATP (eATP) has been reported to be involved in plant growth as a primary messenger in the apoplast. Here, roots of Arabidopsis thaliana seedlings growing in jointed medium bent upon contact with ATP-containing medium to keep away from eATP, showing a marked avoidance response. Roots responded similarly to ADP and bz-ATP but did not respond to AMP and GTP. The eATP avoidance response was reduced in loss-of-function mutants of heterotrimeric G protein α subunit (Gα) (gpa1-1 and gpa1-2) and enhanced in Gα-over-expression (OE) lines (wGα and cGα). Ethylenebis(oxyethylenenitrilo) tetraacetic acid (EGTA) and Gd3+ remarkably suppressed eATP-induced root bending. ATP-stimulated Ca2+ influx was impaired in Gα null mutants and increased in its OE lines. DR5-GFP and PIN2 were asymmetrically distributed in ATP-stimulated root tips, this effect was strongly suppressed by EGTA and diminished in Gα null mutants. In addition, some eATP-induced genes' expression was also impaired in Gα null mutants. Based on these results, we propose that heterotrimeric Gα-regulated Ca2+ influx and PIN2 distribution may be key signaling events in eATP sensing and avoidance response in Arabidopsis thaliana roots.
Extracellular ATP (eATP) plays multiple roles in plant growth and development, and stress responses. It has been revealed that eATP suppresses growth and alters the growth orientation of the root and hypocotyl of Arabidopsis thaliana by affecting auxin transport and localization in these organs. However, the mechanism of the eATP-stimulated auxin distribution remains elusive. Annexins are involved in multiple aspects of plant cellular metabolism, while their role in response to apoplastic signals remains unclear. Here, by using the loss-of-function mutations, we investigated the role of AtANN3 in the eATP-regulated root and hypocotyl growth. Firstly, the inhibitory effects of eATP on root and hypocotyl elongation were weakened or impaired in the AtANN3 null mutants (atann3–1 and atann3–2). Meanwhile, the distribution of DR5-GUS and DR5-GFP indicated that the eATP-induced asymmetric distribution of auxin in the root tips or hypocotyl cells occurred in wild-type control plants, while in atann3–1 mutant seedlings, it was not observed. Further, the eATP-induced asymmetric distribution of PIN2-GFP in root-tip cells or that of PIN3-GFP in hypocotyl cells was reduced in atann3–1 seedlings. Finally, the eATP-induced asymmetric distribution of cytoplasmic vesicles in root-tip cells was impaired in atann3–1 seedlings. Based on these results, we suggest that AtANN3 may be involved in eATP-regulated seedling growth by regulating the distribution of auxin and auxin transporters in vegetative organs.
Extracellular ATP (eATP) exists in the apoplast and plays multiple roles in growth, development, and stress responses. eATP has been revealed to suppresses growth rate and alters growth orientation of root and hypocotyl of Arabidopsis thalianaseedlings by affecting auxin transport in these organs. However, the mechanism of eATP-stimulated auxin distribution remains unclear. Annexins are involved in multiple aspects of plant cellular metabolism, while the role of annexins in response to apoplast signal remains unclear. Here, by using loss-of-function mutants, we investigated the role of several annexins in eATP-regulated root and hypocotyl growth. Since mutants of AtANN3 did not respond to eATP sensitively, the role of AtANN3 in eATP regulated auxin transport was intensively investigated. Firstly, the inhibitory effect of eATP on root or hypocotyl elongation was weakened or impaired in AtANN3 null mutants (atann3-1 and atann3-2). Meanwhile, single-, double- or triple-null mutant of AtANN1, AtANN2or AtANN4 responded to eATP in same manner and degree with Col-0. The distribution of DR5-GUS and DR5-GFP indicated that eATP-induced asymmetric distribution of auxin in root tip or hypocotyl cells, which appeared in wild type controls, were lacking in atann3-1seedlings. Further, eATP-induced asymmetric distribution of PIN2-GFP in root tip cells or PIN3-GFP in hypocotyl cells were reduced in atann3-1 seedlings. Based on these results, we suggest that AtANN3 may be involved in eATP-regulated seedling growth through regulating auxin transport in vegetative organs.
Background Extracellular ATP (eATP) exists in the apoplast of plants and plays multiple roles in growth, development, and stress responses. It has been reported that eATP stimulation suppresses growth rate and alters growth orientation of root and hypocotyls of Arabidopsis thaliana seedlings by affecting auxin accumulation and transport in these organs. However, the mechanism of eATP-stimulated vegetative organ growth remains unclear. Annexins are involved in multiple aspects of plant cellular metabolism, while the role of annexins in response to apoplast signal remains unclear. Here, by using loss-of-function mutants, we investigated the role of several annexins in eATP-regulated root and hypocotyl growth. Since mutant of AtANN3 did not respond to eATP sensitively, the role of AtANN3 in eATP regulated auxin transport was intensively investigated. Results First, the inhibitory effect of eATP on root or hypocotyl elongation was weakened or impaired in AtANN3 null mutants (atann3). Meanwhile, single-, double- or triple-null mutant of AtANN1, AtANN2 or AtANN4 responded to eATP stimulation in same manner and degree with Col-0. The abundance and distribution of Dr5-GUS and Dr5-GFP indicated that eATP-induced accumulation and asymmetric distribution of auxin in root tip or hypocotyl cells, which appeared in wild type controls, were lacking in atann3 seedlings. Further, eATP-induced accumulation and asymmetric distribution of PIN2-GFP in root tip cells or PIN3-GFP in hypocotyl cells were reduced in atann3 seedlings. Conclusions AtANN3 may be involved in eATP-regulated seedling growth through regulating auxin transport and accumulation in vegetative organs.
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