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An Arabidopsis mitochondria-localized RRL protein mediates abscisic acid signal transduction through mitochondrial retrograde regulation involving ABI4.

Yao X, Li J, Liu J, Liu K - J. Exp. Bot. (2015)

Bottom Line: High expression levels of RRL were found in germinating seeds and developing seedlings, as revealed by β-glucuronidase (GUS) staining of ProRRL-GUS transgenic lines.The analyses of the structure and function of mitochondria in the knockout rrl mutant showed that the disruption of RRL causes extensively internally vacuolated mitochondria and reduced ABA-stimulated reactive oxygen species (ROS) production.Furthermore, the results revealed that ABI4 is a downstream regulatory factor in RRL-mediated ABA signalling in seed germination and seedling growth.

View Article: PubMed Central - PubMed

Affiliation: National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan 430070, China.

No MeSH data available.


ABA-insensitive phenotype of the rrl knockout mutant in seed germination and seedling growth. (A) Schematic drawing (not to scale) of T-DNA insertion sites in the loss-of-function mutant allele of the RRL gene (At5G13610). Grey boxes and lines represent exons and introns, respectively. Arrowheads indicate orientations of T-DNA inserts. (LB, left border primer for T-DNA insertion; ATG, translation start codon; TGA, translation stop codon). (B) Semi-quantitative RT-PCR analysis of RRL gene expression in 5-day-old seedlings of Col-0 (wild type), the rrl mutant, and the Com-2 line (complementation line). ACTIN2 (AT3G18780) was used as the internal control. (C) Seeds were sown on MS medium supplemented or not with 1.0 μM ABA. Photographs were taken at day 5 after stratification. Scale bar=0.5cm. (D) Seeds were sown on MS medium supplemented with 0, 0.5, 1.0, 1.5, or 2.0 μM ABA. Germination rates (%) were analysed at day 5 after stratification. Values are the means ±SE of three independent experiments (n=100). *P<0.05 compared with Col-0 control. (E) Seedlings grew for 10 d after transfer from MS medium to MS medium supplemented with 0, 5, 10, 20, or 40 μM ABA. Seedlings were transferred from ABA-free medium to ABA-containing medium 48h after stratification. Scale bar=1cm. (F) Primary root lengths were assayed at day 10 after transfer. Values are the means ±SE of three independent experiments (n=50). *P<0.05 compared with Col-0 control. (This figure is available in colour at JXB online.)
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Figure 2: ABA-insensitive phenotype of the rrl knockout mutant in seed germination and seedling growth. (A) Schematic drawing (not to scale) of T-DNA insertion sites in the loss-of-function mutant allele of the RRL gene (At5G13610). Grey boxes and lines represent exons and introns, respectively. Arrowheads indicate orientations of T-DNA inserts. (LB, left border primer for T-DNA insertion; ATG, translation start codon; TGA, translation stop codon). (B) Semi-quantitative RT-PCR analysis of RRL gene expression in 5-day-old seedlings of Col-0 (wild type), the rrl mutant, and the Com-2 line (complementation line). ACTIN2 (AT3G18780) was used as the internal control. (C) Seeds were sown on MS medium supplemented or not with 1.0 μM ABA. Photographs were taken at day 5 after stratification. Scale bar=0.5cm. (D) Seeds were sown on MS medium supplemented with 0, 0.5, 1.0, 1.5, or 2.0 μM ABA. Germination rates (%) were analysed at day 5 after stratification. Values are the means ±SE of three independent experiments (n=100). *P<0.05 compared with Col-0 control. (E) Seedlings grew for 10 d after transfer from MS medium to MS medium supplemented with 0, 5, 10, 20, or 40 μM ABA. Seedlings were transferred from ABA-free medium to ABA-containing medium 48h after stratification. Scale bar=1cm. (F) Primary root lengths were assayed at day 10 after transfer. Values are the means ±SE of three independent experiments (n=50). *P<0.05 compared with Col-0 control. (This figure is available in colour at JXB online.)

Mentions: To test if this gene has a similar function to RRG, a mutant (SALK_022878C) with the T-DNA inserted in the sixth exon of the RRL gene (+1492) was obtained from the ABRC (Fig. 2A). At5G13610 transcripts were detected in the wild type (Col-0) but not in the rrl mutant (Fig. 2B), suggesting that rrl is a mutant. The rrl mutants showed normal root growth like the wild type (Col-0) plants (see Supplementary Fig. S2 at JXB online), which is different from the retarded root growth of rrg mutants. Moreover, introduction of RRL cDNA driven by the RRG promoter could not recover the retarded root growth of the rrg mutant in complementation assays (see Supplementary Fig. S2). These findings indicated that RRL has different roles in mitochondria in Arabidopsis.


An Arabidopsis mitochondria-localized RRL protein mediates abscisic acid signal transduction through mitochondrial retrograde regulation involving ABI4.

Yao X, Li J, Liu J, Liu K - J. Exp. Bot. (2015)

ABA-insensitive phenotype of the rrl knockout mutant in seed germination and seedling growth. (A) Schematic drawing (not to scale) of T-DNA insertion sites in the loss-of-function mutant allele of the RRL gene (At5G13610). Grey boxes and lines represent exons and introns, respectively. Arrowheads indicate orientations of T-DNA inserts. (LB, left border primer for T-DNA insertion; ATG, translation start codon; TGA, translation stop codon). (B) Semi-quantitative RT-PCR analysis of RRL gene expression in 5-day-old seedlings of Col-0 (wild type), the rrl mutant, and the Com-2 line (complementation line). ACTIN2 (AT3G18780) was used as the internal control. (C) Seeds were sown on MS medium supplemented or not with 1.0 μM ABA. Photographs were taken at day 5 after stratification. Scale bar=0.5cm. (D) Seeds were sown on MS medium supplemented with 0, 0.5, 1.0, 1.5, or 2.0 μM ABA. Germination rates (%) were analysed at day 5 after stratification. Values are the means ±SE of three independent experiments (n=100). *P<0.05 compared with Col-0 control. (E) Seedlings grew for 10 d after transfer from MS medium to MS medium supplemented with 0, 5, 10, 20, or 40 μM ABA. Seedlings were transferred from ABA-free medium to ABA-containing medium 48h after stratification. Scale bar=1cm. (F) Primary root lengths were assayed at day 10 after transfer. Values are the means ±SE of three independent experiments (n=50). *P<0.05 compared with Col-0 control. (This figure is available in colour at JXB online.)
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Figure 2: ABA-insensitive phenotype of the rrl knockout mutant in seed germination and seedling growth. (A) Schematic drawing (not to scale) of T-DNA insertion sites in the loss-of-function mutant allele of the RRL gene (At5G13610). Grey boxes and lines represent exons and introns, respectively. Arrowheads indicate orientations of T-DNA inserts. (LB, left border primer for T-DNA insertion; ATG, translation start codon; TGA, translation stop codon). (B) Semi-quantitative RT-PCR analysis of RRL gene expression in 5-day-old seedlings of Col-0 (wild type), the rrl mutant, and the Com-2 line (complementation line). ACTIN2 (AT3G18780) was used as the internal control. (C) Seeds were sown on MS medium supplemented or not with 1.0 μM ABA. Photographs were taken at day 5 after stratification. Scale bar=0.5cm. (D) Seeds were sown on MS medium supplemented with 0, 0.5, 1.0, 1.5, or 2.0 μM ABA. Germination rates (%) were analysed at day 5 after stratification. Values are the means ±SE of three independent experiments (n=100). *P<0.05 compared with Col-0 control. (E) Seedlings grew for 10 d after transfer from MS medium to MS medium supplemented with 0, 5, 10, 20, or 40 μM ABA. Seedlings were transferred from ABA-free medium to ABA-containing medium 48h after stratification. Scale bar=1cm. (F) Primary root lengths were assayed at day 10 after transfer. Values are the means ±SE of three independent experiments (n=50). *P<0.05 compared with Col-0 control. (This figure is available in colour at JXB online.)
Mentions: To test if this gene has a similar function to RRG, a mutant (SALK_022878C) with the T-DNA inserted in the sixth exon of the RRL gene (+1492) was obtained from the ABRC (Fig. 2A). At5G13610 transcripts were detected in the wild type (Col-0) but not in the rrl mutant (Fig. 2B), suggesting that rrl is a mutant. The rrl mutants showed normal root growth like the wild type (Col-0) plants (see Supplementary Fig. S2 at JXB online), which is different from the retarded root growth of rrg mutants. Moreover, introduction of RRL cDNA driven by the RRG promoter could not recover the retarded root growth of the rrg mutant in complementation assays (see Supplementary Fig. S2). These findings indicated that RRL has different roles in mitochondria in Arabidopsis.

Bottom Line: High expression levels of RRL were found in germinating seeds and developing seedlings, as revealed by β-glucuronidase (GUS) staining of ProRRL-GUS transgenic lines.The analyses of the structure and function of mitochondria in the knockout rrl mutant showed that the disruption of RRL causes extensively internally vacuolated mitochondria and reduced ABA-stimulated reactive oxygen species (ROS) production.Furthermore, the results revealed that ABI4 is a downstream regulatory factor in RRL-mediated ABA signalling in seed germination and seedling growth.

View Article: PubMed Central - PubMed

Affiliation: National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan 430070, China.

No MeSH data available.