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Multiple roles of the transcription factor AtMYBR1/AtMYB44 in ABA signaling, stress responses, and leaf senescence.

Jaradat MR, Feurtado JA, Huang D, Lu Y, Cutler AJ - BMC Plant Biol. (2013)

Bottom Line: MYBR1 appears to exhibit partially redundant functions with AtMYBR2 (MYB77) and double mybr1 X mybr2 mutants exhibited stronger senescence and stress related phenotypes than single mybr1 and mybr2 mutants.MYBR1 is a negative regulator of ABA, stress, wounding responses and blocks senescence.It appears to have a homeostatic function to maintain growth processes in the event of physical damage or stress.

View Article: PubMed Central - HTML - PubMed

Affiliation: Plant Biotechnology Institute, National Research Council of Canada, 110 Gymnasium Place, Saskatoon S7N 0W9, Canada. adrian.cutler@nrc-cnrc.gc.ca.

ABSTRACT

Background: The transcription factor AtMYBR1 (MYB44) is a member of the MYB family of transcription factors and is expressed throughout the plant life cycle and especially in senescing and wounded leaves. It has previously been shown to be involved in responses to abiotic stress and is regulated by phosphorylation.

Results: When MYBR1 was over-expressed under the control of the constitutive 35S promoter in Arabidopsis thaliana (OxMYBR1), leaf senescence was delayed. In contrast loss-of-function mybr1 plants showed more rapid chlorophyll loss and senescence. The MYBR1 promoter strongly drove β-GLUCURONIDASE reporter gene expression in tissues immediately after wounding and many wounding/pathogenesis genes were downregulated in OxMYBR1. OxMYBR1 plants were more susceptible to injury under water stress than wildtype, which was correlated with suppression of many ABA inducible stress genes in OxMYBR1. Conversely, mybr1 plants were more tolerant of water stress and exhibited reduced rates of water loss from leaves. MYBR1 physically interacted with ABA receptor PYR1-LIKE8 (PYL8) suggesting a direct involvement of MYBR1 in early ABA signaling. MYBR1 appears to exhibit partially redundant functions with AtMYBR2 (MYB77) and double mybr1 X mybr2 mutants exhibited stronger senescence and stress related phenotypes than single mybr1 and mybr2 mutants.

Conclusions: MYBR1 is a negative regulator of ABA, stress, wounding responses and blocks senescence. It appears to have a homeostatic function to maintain growth processes in the event of physical damage or stress.

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Effect of MYBR1 on leaf senescence in a detached leaf assay. (A) True leaves numbers 3–6 were harvested from 30 d old soil grown plants and incubated on filter paper wetted with 3 mM MES buffer (pH 5.7). Leaves 3 and 4 were photographed after 6 d treatment and leaves 5 and 6 were photographed after 7 d treatment. Leaves from OxMYBR1 plants of three independent lines (#31-3, 1–7 and 42–6) showed delayed senescence relative to other genotypes and leaves from mybr1 and double mutant plants exhibited premature leaf senescence relative to leaves from WT (leaves 5 and 6) and OxMYBR1 plants (all leaves). Two sets of experiment were carried out as above (A) in four replicates and 12 plants in each replicate. Statistical significance was determined using one-way ANOVA with Tukey using the statistical software ‘R’ (P < 0.05). (B) In one set of experiments, chlorophyll was extracted and measured on 0 d. The chlorophyll content was higher in one line of OxMYBR1 (#42-6) and reciprocal double mutants of mybr1 and mybr2 than other genotypes. (C) In another set, chlorophyll was extracted and measured on 6 d for leaves 3–4 and on 7d for leaves 5–6 and the percentage chlorophyll retention was calculated relative to 0 d from (B). Chlorophyll retention was generally higher in OxMYBR1 genotypes. (D) Using three independent homozygous MYBR1pro:GUS lines (#5-1, 7–6 and X1-4), experiments were carried out as above (A) in two replicates. All leaves from each plant were harvested. GUS staining was performed on 0 d (untreated) and after 4 d of dark induced senescence. Asterisks indicate yellow leaves before and after GUS staining. After the treatment, GUS staining was higher in senescent leaves, but in green leaves was lower than corresponding control leaves.
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Figure 5: Effect of MYBR1 on leaf senescence in a detached leaf assay. (A) True leaves numbers 3–6 were harvested from 30 d old soil grown plants and incubated on filter paper wetted with 3 mM MES buffer (pH 5.7). Leaves 3 and 4 were photographed after 6 d treatment and leaves 5 and 6 were photographed after 7 d treatment. Leaves from OxMYBR1 plants of three independent lines (#31-3, 1–7 and 42–6) showed delayed senescence relative to other genotypes and leaves from mybr1 and double mutant plants exhibited premature leaf senescence relative to leaves from WT (leaves 5 and 6) and OxMYBR1 plants (all leaves). Two sets of experiment were carried out as above (A) in four replicates and 12 plants in each replicate. Statistical significance was determined using one-way ANOVA with Tukey using the statistical software ‘R’ (P < 0.05). (B) In one set of experiments, chlorophyll was extracted and measured on 0 d. The chlorophyll content was higher in one line of OxMYBR1 (#42-6) and reciprocal double mutants of mybr1 and mybr2 than other genotypes. (C) In another set, chlorophyll was extracted and measured on 6 d for leaves 3–4 and on 7d for leaves 5–6 and the percentage chlorophyll retention was calculated relative to 0 d from (B). Chlorophyll retention was generally higher in OxMYBR1 genotypes. (D) Using three independent homozygous MYBR1pro:GUS lines (#5-1, 7–6 and X1-4), experiments were carried out as above (A) in two replicates. All leaves from each plant were harvested. GUS staining was performed on 0 d (untreated) and after 4 d of dark induced senescence. Asterisks indicate yellow leaves before and after GUS staining. After the treatment, GUS staining was higher in senescent leaves, but in green leaves was lower than corresponding control leaves.

Mentions: Early in leaf senescence chloroplasts disassemble with subsequent degradation of chlorophyll and visible leaf yellowing. To further investigate the role of MYBR1 in leaf longevity, detached rosette true leaves numbers 3–6 (counted by order of emergence), from 30 d old soil grown plants were incubated in buffer as described [26] in two different sets. Leaves were photographed and the chlorophyll content was quantified on 0 d for one set and after 6–7 d of dark-induced senescence treatment for the other set. In freshly harvested leaves, the chlorophyll content was higher in one line of OxMYBR1 (#42-6) and two reciprocal double mutants of mybr1 and mybr2 than the rest of the genotypes (Figure 5B). Following dark-induced senescence, OxMYBR1 lines showed increased leaf longevity and slowed chlorophyll degradation relative to WT leaves (Figure 5A and 5C). Interestingly, increased leaf longevity in OxMYBR1 lines was in contrast to that in mybr1 and reciprocal mybr1 & mybr2 mutants which exhibited early leaf senescence and accelerated chlorophyll degradation relative to WT (Figure 5A and 5C) although the chlorophyll content before treatment was relatively high in mybr1 and mybr2 double mutants (Figure 5B). Indeed, among the eight genotypes, leaves of mybr1 & mybr2 were least green following senescence treatment and showed the fastest chlorophyll breakdown (Figure 5A and 5C).


Multiple roles of the transcription factor AtMYBR1/AtMYB44 in ABA signaling, stress responses, and leaf senescence.

Jaradat MR, Feurtado JA, Huang D, Lu Y, Cutler AJ - BMC Plant Biol. (2013)

Effect of MYBR1 on leaf senescence in a detached leaf assay. (A) True leaves numbers 3–6 were harvested from 30 d old soil grown plants and incubated on filter paper wetted with 3 mM MES buffer (pH 5.7). Leaves 3 and 4 were photographed after 6 d treatment and leaves 5 and 6 were photographed after 7 d treatment. Leaves from OxMYBR1 plants of three independent lines (#31-3, 1–7 and 42–6) showed delayed senescence relative to other genotypes and leaves from mybr1 and double mutant plants exhibited premature leaf senescence relative to leaves from WT (leaves 5 and 6) and OxMYBR1 plants (all leaves). Two sets of experiment were carried out as above (A) in four replicates and 12 plants in each replicate. Statistical significance was determined using one-way ANOVA with Tukey using the statistical software ‘R’ (P < 0.05). (B) In one set of experiments, chlorophyll was extracted and measured on 0 d. The chlorophyll content was higher in one line of OxMYBR1 (#42-6) and reciprocal double mutants of mybr1 and mybr2 than other genotypes. (C) In another set, chlorophyll was extracted and measured on 6 d for leaves 3–4 and on 7d for leaves 5–6 and the percentage chlorophyll retention was calculated relative to 0 d from (B). Chlorophyll retention was generally higher in OxMYBR1 genotypes. (D) Using three independent homozygous MYBR1pro:GUS lines (#5-1, 7–6 and X1-4), experiments were carried out as above (A) in two replicates. All leaves from each plant were harvested. GUS staining was performed on 0 d (untreated) and after 4 d of dark induced senescence. Asterisks indicate yellow leaves before and after GUS staining. After the treatment, GUS staining was higher in senescent leaves, but in green leaves was lower than corresponding control leaves.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4219380&req=5

Figure 5: Effect of MYBR1 on leaf senescence in a detached leaf assay. (A) True leaves numbers 3–6 were harvested from 30 d old soil grown plants and incubated on filter paper wetted with 3 mM MES buffer (pH 5.7). Leaves 3 and 4 were photographed after 6 d treatment and leaves 5 and 6 were photographed after 7 d treatment. Leaves from OxMYBR1 plants of three independent lines (#31-3, 1–7 and 42–6) showed delayed senescence relative to other genotypes and leaves from mybr1 and double mutant plants exhibited premature leaf senescence relative to leaves from WT (leaves 5 and 6) and OxMYBR1 plants (all leaves). Two sets of experiment were carried out as above (A) in four replicates and 12 plants in each replicate. Statistical significance was determined using one-way ANOVA with Tukey using the statistical software ‘R’ (P < 0.05). (B) In one set of experiments, chlorophyll was extracted and measured on 0 d. The chlorophyll content was higher in one line of OxMYBR1 (#42-6) and reciprocal double mutants of mybr1 and mybr2 than other genotypes. (C) In another set, chlorophyll was extracted and measured on 6 d for leaves 3–4 and on 7d for leaves 5–6 and the percentage chlorophyll retention was calculated relative to 0 d from (B). Chlorophyll retention was generally higher in OxMYBR1 genotypes. (D) Using three independent homozygous MYBR1pro:GUS lines (#5-1, 7–6 and X1-4), experiments were carried out as above (A) in two replicates. All leaves from each plant were harvested. GUS staining was performed on 0 d (untreated) and after 4 d of dark induced senescence. Asterisks indicate yellow leaves before and after GUS staining. After the treatment, GUS staining was higher in senescent leaves, but in green leaves was lower than corresponding control leaves.
Mentions: Early in leaf senescence chloroplasts disassemble with subsequent degradation of chlorophyll and visible leaf yellowing. To further investigate the role of MYBR1 in leaf longevity, detached rosette true leaves numbers 3–6 (counted by order of emergence), from 30 d old soil grown plants were incubated in buffer as described [26] in two different sets. Leaves were photographed and the chlorophyll content was quantified on 0 d for one set and after 6–7 d of dark-induced senescence treatment for the other set. In freshly harvested leaves, the chlorophyll content was higher in one line of OxMYBR1 (#42-6) and two reciprocal double mutants of mybr1 and mybr2 than the rest of the genotypes (Figure 5B). Following dark-induced senescence, OxMYBR1 lines showed increased leaf longevity and slowed chlorophyll degradation relative to WT leaves (Figure 5A and 5C). Interestingly, increased leaf longevity in OxMYBR1 lines was in contrast to that in mybr1 and reciprocal mybr1 & mybr2 mutants which exhibited early leaf senescence and accelerated chlorophyll degradation relative to WT (Figure 5A and 5C) although the chlorophyll content before treatment was relatively high in mybr1 and mybr2 double mutants (Figure 5B). Indeed, among the eight genotypes, leaves of mybr1 & mybr2 were least green following senescence treatment and showed the fastest chlorophyll breakdown (Figure 5A and 5C).

Bottom Line: MYBR1 appears to exhibit partially redundant functions with AtMYBR2 (MYB77) and double mybr1 X mybr2 mutants exhibited stronger senescence and stress related phenotypes than single mybr1 and mybr2 mutants.MYBR1 is a negative regulator of ABA, stress, wounding responses and blocks senescence.It appears to have a homeostatic function to maintain growth processes in the event of physical damage or stress.

View Article: PubMed Central - HTML - PubMed

Affiliation: Plant Biotechnology Institute, National Research Council of Canada, 110 Gymnasium Place, Saskatoon S7N 0W9, Canada. adrian.cutler@nrc-cnrc.gc.ca.

ABSTRACT

Background: The transcription factor AtMYBR1 (MYB44) is a member of the MYB family of transcription factors and is expressed throughout the plant life cycle and especially in senescing and wounded leaves. It has previously been shown to be involved in responses to abiotic stress and is regulated by phosphorylation.

Results: When MYBR1 was over-expressed under the control of the constitutive 35S promoter in Arabidopsis thaliana (OxMYBR1), leaf senescence was delayed. In contrast loss-of-function mybr1 plants showed more rapid chlorophyll loss and senescence. The MYBR1 promoter strongly drove β-GLUCURONIDASE reporter gene expression in tissues immediately after wounding and many wounding/pathogenesis genes were downregulated in OxMYBR1. OxMYBR1 plants were more susceptible to injury under water stress than wildtype, which was correlated with suppression of many ABA inducible stress genes in OxMYBR1. Conversely, mybr1 plants were more tolerant of water stress and exhibited reduced rates of water loss from leaves. MYBR1 physically interacted with ABA receptor PYR1-LIKE8 (PYL8) suggesting a direct involvement of MYBR1 in early ABA signaling. MYBR1 appears to exhibit partially redundant functions with AtMYBR2 (MYB77) and double mybr1 X mybr2 mutants exhibited stronger senescence and stress related phenotypes than single mybr1 and mybr2 mutants.

Conclusions: MYBR1 is a negative regulator of ABA, stress, wounding responses and blocks senescence. It appears to have a homeostatic function to maintain growth processes in the event of physical damage or stress.

Show MeSH
Related in: MedlinePlus