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The Arabidopsis a zinc finger domain protein ARS1 is essential for seed germination and ROS homeostasis in response to ABA and oxidative stress

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ABSTRACT

The phytohormone abscisic acid (ABA) induces accumulation of reactive oxygen species (ROS), which can disrupt seed dormancy and plant development. Here, we report the isolation and characterization of an Arabidopsis thaliana mutant called ars1 (aba and ros sensitive 1) that showed hypersensitivity to ABA during seed germination and to methyl viologen (MV) at the seedling stage. ARS1 encodes a nuclear protein with one zinc finger domain, two nuclear localization signal (NLS) domains, and one nuclear export signal (NES). The ars1 mutants showed reduced expression of a gene for superoxide dismutase (CSD3) and enhanced accumulation of ROS after ABA treatment. Transient expression of ARS1 in Arabidopsis protoplasts strongly suppressed ABA-mediated ROS production. Interestingly, nuclear-localized ARS1 translocated to the cytoplasm in response to treatment with ABA, H2O2, or MV. Taken together, these results suggest that ARS1 modulates seed germination and ROS homeostasis in response to ABA and oxidative stress in plants.

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ARS1 is translocated from the nucleus to the cytoplasm in response to ABA and oxidative stresses. (A) Protoplasts were isolated from leaves of 3-week-old WT Col-0 plants transformed with ARS1::sGFP. Twelve hours after transformation, protoplasts were treated with ABA (100 μM), H2O2 (1.5 mM) or MV (3 μM) for the indicated times. Bars indicate 20 μm. (B) Percentage of ARS1 signal exported to the cytoplasm in response to ABA, H2O2, and MV shown in (A). Bars represent mean ± SD of three biological replicates with three technical replicates each (n = 200). Asterisks represent significant differences from the 0 h(∗∗p-value ≤0.01, Student’s t-test).
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Figure 6: ARS1 is translocated from the nucleus to the cytoplasm in response to ABA and oxidative stresses. (A) Protoplasts were isolated from leaves of 3-week-old WT Col-0 plants transformed with ARS1::sGFP. Twelve hours after transformation, protoplasts were treated with ABA (100 μM), H2O2 (1.5 mM) or MV (3 μM) for the indicated times. Bars indicate 20 μm. (B) Percentage of ARS1 signal exported to the cytoplasm in response to ABA, H2O2, and MV shown in (A). Bars represent mean ± SD of three biological replicates with three technical replicates each (n = 200). Asterisks represent significant differences from the 0 h(∗∗p-value ≤0.01, Student’s t-test).

Mentions: Protein sequence analysis revealed that ARS1 has a putative NES motif at its C terminus (Figure 2B). The presence of the putative NES signal suggested that ARS1 could be exported from the nucleus in response to certain stress conditions and led us to investigate the changes of subcellular localization of ARS1 in Arabidopsis cells. To analyze the subcellular localization of ARS1 under stress conditions, we first transiently expressed ARS1::sGFP in Arabidopsis (Col-0) protoplast cells. Twelve hours after transformation of ARS1::sGFP into protoplasts, we treated the protoplasts with ABA, H2O2, or methyl viologen (MV) as ROS triggers. In the absence of stressors, ARS1 clearly localized to the nucleus (Figures 2C and 6A, Supplementary Figure S5). Surprisingly, nuclear-localized ARS1 was abundantly translocated to the cytoplasm in response to ABA, H2O2, and MV. (Figure 6, Supplementary Figure S6). These translocation patterns of ARS1 in protoplasts markedly increased as the stress duration increased up to 5 h (Figure 6B). These results indicate that the putative NES motif is likely important for ARS1 function in stress responses. Together, this evidence suggests that ARS1 exists as an inactive form in the nucleus, but changes its localization to the cytoplasm as a result of ROS signals induced by ABA and other stresses to repress ABA/stress-induced ROS production in plant cells.


The Arabidopsis a zinc finger domain protein ARS1 is essential for seed germination and ROS homeostasis in response to ABA and oxidative stress
ARS1 is translocated from the nucleus to the cytoplasm in response to ABA and oxidative stresses. (A) Protoplasts were isolated from leaves of 3-week-old WT Col-0 plants transformed with ARS1::sGFP. Twelve hours after transformation, protoplasts were treated with ABA (100 μM), H2O2 (1.5 mM) or MV (3 μM) for the indicated times. Bars indicate 20 μm. (B) Percentage of ARS1 signal exported to the cytoplasm in response to ABA, H2O2, and MV shown in (A). Bars represent mean ± SD of three biological replicates with three technical replicates each (n = 200). Asterisks represent significant differences from the 0 h(∗∗p-value ≤0.01, Student’s t-test).
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Figure 6: ARS1 is translocated from the nucleus to the cytoplasm in response to ABA and oxidative stresses. (A) Protoplasts were isolated from leaves of 3-week-old WT Col-0 plants transformed with ARS1::sGFP. Twelve hours after transformation, protoplasts were treated with ABA (100 μM), H2O2 (1.5 mM) or MV (3 μM) for the indicated times. Bars indicate 20 μm. (B) Percentage of ARS1 signal exported to the cytoplasm in response to ABA, H2O2, and MV shown in (A). Bars represent mean ± SD of three biological replicates with three technical replicates each (n = 200). Asterisks represent significant differences from the 0 h(∗∗p-value ≤0.01, Student’s t-test).
Mentions: Protein sequence analysis revealed that ARS1 has a putative NES motif at its C terminus (Figure 2B). The presence of the putative NES signal suggested that ARS1 could be exported from the nucleus in response to certain stress conditions and led us to investigate the changes of subcellular localization of ARS1 in Arabidopsis cells. To analyze the subcellular localization of ARS1 under stress conditions, we first transiently expressed ARS1::sGFP in Arabidopsis (Col-0) protoplast cells. Twelve hours after transformation of ARS1::sGFP into protoplasts, we treated the protoplasts with ABA, H2O2, or methyl viologen (MV) as ROS triggers. In the absence of stressors, ARS1 clearly localized to the nucleus (Figures 2C and 6A, Supplementary Figure S5). Surprisingly, nuclear-localized ARS1 was abundantly translocated to the cytoplasm in response to ABA, H2O2, and MV. (Figure 6, Supplementary Figure S6). These translocation patterns of ARS1 in protoplasts markedly increased as the stress duration increased up to 5 h (Figure 6B). These results indicate that the putative NES motif is likely important for ARS1 function in stress responses. Together, this evidence suggests that ARS1 exists as an inactive form in the nucleus, but changes its localization to the cytoplasm as a result of ROS signals induced by ABA and other stresses to repress ABA/stress-induced ROS production in plant cells.

View Article: PubMed Central

ABSTRACT

The phytohormone abscisic acid (ABA) induces accumulation of reactive oxygen species (ROS), which can disrupt seed dormancy and plant development. Here, we report the isolation and characterization of an Arabidopsis thaliana mutant called ars1 (aba and ros sensitive 1) that showed hypersensitivity to ABA during seed germination and to methyl viologen (MV) at the seedling stage. ARS1 encodes a nuclear protein with one zinc finger domain, two nuclear localization signal (NLS) domains, and one nuclear export signal (NES). The ars1 mutants showed reduced expression of a gene for superoxide dismutase (CSD3) and enhanced accumulation of ROS after ABA treatment. Transient expression of ARS1 in Arabidopsis protoplasts strongly suppressed ABA-mediated ROS production. Interestingly, nuclear-localized ARS1 translocated to the cytoplasm in response to treatment with ABA, H2O2, or MV. Taken together, these results suggest that ARS1 modulates seed germination and ROS homeostasis in response to ABA and oxidative stress in plants.

No MeSH data available.


Related in: MedlinePlus