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AKIN10 delays flowering by inactivating IDD8 transcription factor through protein phosphorylation in Arabidopsis.

Jeong EY, Seo PJ, Woo JC, Park CM - BMC Plant Biol. (2015)

Bottom Line: We found that AKIN10 interacts with IDD8 in the nucleus.AKIN10-mediated phosphorylation did not affect the subcellular localization and DNA-binding property of IDD8.Instead, the transcriptional activation activity of the phosphorylated IDD8 was significantly reduced.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Seoul National University, Seoul, 151-742, South Korea. eyjeong@snu.ac.kr.

ABSTRACT

Background: Sugar plays a central role as a source of carbon metabolism and energy production and a signaling molecule in diverse growth and developmental processes and environmental adaptation in plants. It is known that sugar metabolism and allocation between different physiological functions is intimately associated with flowering transition in many plant species. The INDETERMINATE DOMAIN (IDD)-containing transcription factor IDD8 regulates flowering time by modulating sugar metabolism and transport under sugar-limiting conditions in Arabidopsis. Meanwhile, it has been reported that SUCROSE NONFERMENTING-1-RELATED PROTEIN KINASE 1 (SnRK1), which acts as a sensor of cellular energy metabolism, is activated by sugar deprivation. Notably, SnRK1-overexpressing plants and IDD8-deficient mutants exhibit similar phenotypes, including delayed flowering, suggesting that SnRK1 is involved in the IDD8-mediated metabolic control of flowering.

Results: We examined whether the sugar deprivation-sensing SnRK1 is functionally associated with IDD8 in flowering time control through biochemical and molecular genetic approaches. Overproduction of AKIN10, the catalytic subunit of SnRK1, delayed flowering in Arabidopsis, as was observed in IDD8-deficient idd8-3 mutant. We found that AKIN10 interacts with IDD8 in the nucleus. Consequently, AKIN10 phosphorylates IDD8 primarily at two serine (Ser) residues, Ser-178 and Ser-182, which reside in the fourth zinc finger (ZF) domain that mediates DNA binding and protein-protein interactions. AKIN10-mediated phosphorylation did not affect the subcellular localization and DNA-binding property of IDD8. Instead, the transcriptional activation activity of the phosphorylated IDD8 was significantly reduced. Together, these observations indicate that AKIN10 antagonizes the IDD8 function in flowering time control, a notion that is consistent with the delayed flowering phenotypes of AKIN10-overexpressing plants and idd8-3 mutant.

Conclusion: Our data show that SnRK1 and its substrate IDD8 constitute a sugar metabolic pathway that mediates the timing of flowering under sugar deprivation conditions. In this signaling scheme, the SnRK1 signals are directly integrated into the IDD8-mediated gene regulatory network that governs flowering transition in response to fluctuations in sugar metabolism, further supporting the metabolic control of flowering.

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Related in: MedlinePlus

Phosphorylation of IDD8 by AKIN10. The in vitro phosphorylation assays were conducted using recombinant GST-AKIN10 and GST-AKIN11 fusion proteins and MBP-IDD8 fusion protein prepared in E. coli cells (upper panel). Part of Coomassie Blue-stained gel was displayed as a loading control (lower panel). kDa, kilodalton.
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Fig3: Phosphorylation of IDD8 by AKIN10. The in vitro phosphorylation assays were conducted using recombinant GST-AKIN10 and GST-AKIN11 fusion proteins and MBP-IDD8 fusion protein prepared in E. coli cells (upper panel). Part of Coomassie Blue-stained gel was displayed as a loading control (lower panel). kDa, kilodalton.

Mentions: We produced recombinant maltose-binding protein-IDD8 (MBP-IDD8) and GST-AKIN fusion proteins in E.coli cells, which were purified by affinity chromatography and immunologically quantified (Additional file 5A). The in vitro kinase assays showed that AKIN10 possesses an autophosphorylation activity, while AKIN11 does not (Figure 3). It was also evident that AKIN10, but not AKIN11, phosphorylates IDD8. Although both 10-ox and 11-ox plants exhibited delayed flowering (Figure 1) and IDD8 interacts with both AKIN10 and AKIN11, IDD8 may not be directly targeted by AKIN11 at least in controlling flowering time.Figure 3


AKIN10 delays flowering by inactivating IDD8 transcription factor through protein phosphorylation in Arabidopsis.

Jeong EY, Seo PJ, Woo JC, Park CM - BMC Plant Biol. (2015)

Phosphorylation of IDD8 by AKIN10. The in vitro phosphorylation assays were conducted using recombinant GST-AKIN10 and GST-AKIN11 fusion proteins and MBP-IDD8 fusion protein prepared in E. coli cells (upper panel). Part of Coomassie Blue-stained gel was displayed as a loading control (lower panel). kDa, kilodalton.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4416337&req=5

Fig3: Phosphorylation of IDD8 by AKIN10. The in vitro phosphorylation assays were conducted using recombinant GST-AKIN10 and GST-AKIN11 fusion proteins and MBP-IDD8 fusion protein prepared in E. coli cells (upper panel). Part of Coomassie Blue-stained gel was displayed as a loading control (lower panel). kDa, kilodalton.
Mentions: We produced recombinant maltose-binding protein-IDD8 (MBP-IDD8) and GST-AKIN fusion proteins in E.coli cells, which were purified by affinity chromatography and immunologically quantified (Additional file 5A). The in vitro kinase assays showed that AKIN10 possesses an autophosphorylation activity, while AKIN11 does not (Figure 3). It was also evident that AKIN10, but not AKIN11, phosphorylates IDD8. Although both 10-ox and 11-ox plants exhibited delayed flowering (Figure 1) and IDD8 interacts with both AKIN10 and AKIN11, IDD8 may not be directly targeted by AKIN11 at least in controlling flowering time.Figure 3

Bottom Line: We found that AKIN10 interacts with IDD8 in the nucleus.AKIN10-mediated phosphorylation did not affect the subcellular localization and DNA-binding property of IDD8.Instead, the transcriptional activation activity of the phosphorylated IDD8 was significantly reduced.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Seoul National University, Seoul, 151-742, South Korea. eyjeong@snu.ac.kr.

ABSTRACT

Background: Sugar plays a central role as a source of carbon metabolism and energy production and a signaling molecule in diverse growth and developmental processes and environmental adaptation in plants. It is known that sugar metabolism and allocation between different physiological functions is intimately associated with flowering transition in many plant species. The INDETERMINATE DOMAIN (IDD)-containing transcription factor IDD8 regulates flowering time by modulating sugar metabolism and transport under sugar-limiting conditions in Arabidopsis. Meanwhile, it has been reported that SUCROSE NONFERMENTING-1-RELATED PROTEIN KINASE 1 (SnRK1), which acts as a sensor of cellular energy metabolism, is activated by sugar deprivation. Notably, SnRK1-overexpressing plants and IDD8-deficient mutants exhibit similar phenotypes, including delayed flowering, suggesting that SnRK1 is involved in the IDD8-mediated metabolic control of flowering.

Results: We examined whether the sugar deprivation-sensing SnRK1 is functionally associated with IDD8 in flowering time control through biochemical and molecular genetic approaches. Overproduction of AKIN10, the catalytic subunit of SnRK1, delayed flowering in Arabidopsis, as was observed in IDD8-deficient idd8-3 mutant. We found that AKIN10 interacts with IDD8 in the nucleus. Consequently, AKIN10 phosphorylates IDD8 primarily at two serine (Ser) residues, Ser-178 and Ser-182, which reside in the fourth zinc finger (ZF) domain that mediates DNA binding and protein-protein interactions. AKIN10-mediated phosphorylation did not affect the subcellular localization and DNA-binding property of IDD8. Instead, the transcriptional activation activity of the phosphorylated IDD8 was significantly reduced. Together, these observations indicate that AKIN10 antagonizes the IDD8 function in flowering time control, a notion that is consistent with the delayed flowering phenotypes of AKIN10-overexpressing plants and idd8-3 mutant.

Conclusion: Our data show that SnRK1 and its substrate IDD8 constitute a sugar metabolic pathway that mediates the timing of flowering under sugar deprivation conditions. In this signaling scheme, the SnRK1 signals are directly integrated into the IDD8-mediated gene regulatory network that governs flowering transition in response to fluctuations in sugar metabolism, further supporting the metabolic control of flowering.

Show MeSH
Related in: MedlinePlus