Mol. Cells 2015; 38(1): 81-88
Published online November 26, 2014
https://doi.org/10.14348/molcells.2015.2254
© The Korean Society for Molecular and Cellular Biology
Correspondence to : *Correspondence: ncpaek@snu.ac.kr
Flowering time (or heading date) is controlled by intrinsic genetic programs in response to environmental cues, such as photoperiod and temperature. Rice, a facultative short-day (SD) plant, flowers early in SD and late in long-day (LD) conditions. Casein kinases (CKs) generally act as positive regulators in many signaling pathways in plants. In rice,
Keywords casein kinase I, casein kinase 2α, flowering time, OsPRR37, phosphorylation, rice
In plants, complex interactions between endogenous circadian clock components and environmental factors trigger flowering; these environmental factors include seasonal changes in day length (photoperiod) and temperature. To date, research in model systems and crops has identified many regulatory genes controlling flowering time (also known as heading date), for example in the dicot
Cultivated rice is a facultative short-day (SD) plant that flowers early in SD (< 10-h light/day) and late in long day (LD; > 14-h light/day) conditions (Izawa, 2007; Tsuji et al., 2008). Modern rice cultivation spans geographical latitudes from 53°N to 40°S and photoperiod sensitivity affects adaptation for growth at these different latitudes, significantly affecting grain yield (Izawa, 2007). For example,
In
The highly conserved serine/threonine-specific casein kinases (CKs) control various signal transduction processes in eukaryotes (Knippschild et al., 2005; Mulekar and Huq, 2014). In plants, CKI and CK2 affect the regulation of flowering time and circadian rhythm. In
Pseudo-Response Regulators (PRRs) are essential circadian clock components in
Here, we show the direct interactions of Hd6/CK2α and Hd16/CKI with Hd2/PRR37
The cDNAs of
Eluted HisMBP proteins (a negative control) and HisMBPPRR37 were incubated with GST-CK2α or His-CKI in GST-Bind Agarose Resin (Elpis Biotech, Korea) or MBP-Bind Agarose Resin (Elpis Biotech, Korea), respectively, at 4°C for 1 h. The resin was washed four times in GST pull-down washing buffer (50 mM Tris-HCl, pH 7.5, 200 mM NaCl, 0.5 mM β-mercaptoethanol, 1% Triton X-100 and 0.2% glycerol) or MBP pull-down washing buffer (20 mM Tris-HCl, pH 7.4, 200 mM NaCl, 10 mM β-mercaptoethanol, 1 mM EDTA). To boil the proteins bound to the resin, 5x SDS-PAGE loading buffer was added and heated at 100°C for 4 min. Proteins were resolved by SDS-PAGE and immunoblotted using antibodies against GST (Santa Cruz Biotechnology, USA), MBP (Santa Cruz Bio-technology, USA), and His(6x)-tag (Abcam, USA)
For YFP-tagged and partial YFP-tagged PRR37 constructs, PCR-amplified
To do the
The suppression of flowering by
To examine their potential interactions
To confirm the interactions of CK2α-PRR37 and CKI-PRR37
CK2α and CKI can phosphorylate specific substrates; CK2α phosphorylates OsLHY in the circadian clock and flowering regulation (Ogiso et al., 2010), and CKI phosphorylates SLR1 in GA signaling and Ghd7 in flowering regulation (Dai and Xue, 2010; Hori et al., 2013). To examine whether these two protein kinases can phosphorylate PRR37, we conducted
To determine the phosphorylation sites in PRR37, we first examined the PRR37 sequence with a casein kinase-specific phosphorylation site prediction algorithm, KinasePhos 2.0 (
In
In floral repression under non-inductive LD conditions, Hd6/CK2α may phosphorylate downstream LD-dependent floral repressor(s). Several studies have reported the epistatic interactions between
Second,
It is noteworthy that CKI and CK2α phosphorylate distinct regions in the PRR37 protein; CKI phosphorylates the recombinant partial proteins PRR37m and PRR37c (Fig. 4B), but CK2α phosphorylates only PRR37m (Fig. 4B). This suggests that the levels of phosphorylation by CKI and CK2α might separately regulate the activity and/or stability of PRR37. Further
Natural variants of
In conclusion, we propose that CKI and CK2α may contribute to enhancing the photoperiod sensitivity of rice through phosphorylation of PRR37. Furthermore, our results provide new, important insights into CKI and/or CK2-mediated phosphorylation of PRR proteins in other plants including barley, wheat, sorghum, maize, and
Mol. Cells 2015; 38(1): 81-88
Published online January 31, 2015 https://doi.org/10.14348/molcells.2015.2254
Copyright © The Korean Society for Molecular and Cellular Biology.
Choon-Tak Kwon1, Bon-Hyuk Koo1, Dami Kim1, Soo-Cheul Yoo2, and Nam-Chon Paek1,*
1Department of Plant Science, Plant Genomics and Breeding Institute, and Research Institute for Agriculture and Life Sciences, Seoul National University, Seoul 151-921, Korea, 2Department of Bioresource and Rural System of Engineering, Hankyong National University, Ansung 456-749, Korea
Correspondence to:*Correspondence: ncpaek@snu.ac.kr
Flowering time (or heading date) is controlled by intrinsic genetic programs in response to environmental cues, such as photoperiod and temperature. Rice, a facultative short-day (SD) plant, flowers early in SD and late in long-day (LD) conditions. Casein kinases (CKs) generally act as positive regulators in many signaling pathways in plants. In rice,
Keywords: casein kinase I, casein kinase 2α, flowering time, OsPRR37, phosphorylation, rice
In plants, complex interactions between endogenous circadian clock components and environmental factors trigger flowering; these environmental factors include seasonal changes in day length (photoperiod) and temperature. To date, research in model systems and crops has identified many regulatory genes controlling flowering time (also known as heading date), for example in the dicot
Cultivated rice is a facultative short-day (SD) plant that flowers early in SD (< 10-h light/day) and late in long day (LD; > 14-h light/day) conditions (Izawa, 2007; Tsuji et al., 2008). Modern rice cultivation spans geographical latitudes from 53°N to 40°S and photoperiod sensitivity affects adaptation for growth at these different latitudes, significantly affecting grain yield (Izawa, 2007). For example,
In
The highly conserved serine/threonine-specific casein kinases (CKs) control various signal transduction processes in eukaryotes (Knippschild et al., 2005; Mulekar and Huq, 2014). In plants, CKI and CK2 affect the regulation of flowering time and circadian rhythm. In
Pseudo-Response Regulators (PRRs) are essential circadian clock components in
Here, we show the direct interactions of Hd6/CK2α and Hd16/CKI with Hd2/PRR37
The cDNAs of
Eluted HisMBP proteins (a negative control) and HisMBPPRR37 were incubated with GST-CK2α or His-CKI in GST-Bind Agarose Resin (Elpis Biotech, Korea) or MBP-Bind Agarose Resin (Elpis Biotech, Korea), respectively, at 4°C for 1 h. The resin was washed four times in GST pull-down washing buffer (50 mM Tris-HCl, pH 7.5, 200 mM NaCl, 0.5 mM β-mercaptoethanol, 1% Triton X-100 and 0.2% glycerol) or MBP pull-down washing buffer (20 mM Tris-HCl, pH 7.4, 200 mM NaCl, 10 mM β-mercaptoethanol, 1 mM EDTA). To boil the proteins bound to the resin, 5x SDS-PAGE loading buffer was added and heated at 100°C for 4 min. Proteins were resolved by SDS-PAGE and immunoblotted using antibodies against GST (Santa Cruz Biotechnology, USA), MBP (Santa Cruz Bio-technology, USA), and His(6x)-tag (Abcam, USA)
For YFP-tagged and partial YFP-tagged PRR37 constructs, PCR-amplified
To do the
The suppression of flowering by
To examine their potential interactions
To confirm the interactions of CK2α-PRR37 and CKI-PRR37
CK2α and CKI can phosphorylate specific substrates; CK2α phosphorylates OsLHY in the circadian clock and flowering regulation (Ogiso et al., 2010), and CKI phosphorylates SLR1 in GA signaling and Ghd7 in flowering regulation (Dai and Xue, 2010; Hori et al., 2013). To examine whether these two protein kinases can phosphorylate PRR37, we conducted
To determine the phosphorylation sites in PRR37, we first examined the PRR37 sequence with a casein kinase-specific phosphorylation site prediction algorithm, KinasePhos 2.0 (
In
In floral repression under non-inductive LD conditions, Hd6/CK2α may phosphorylate downstream LD-dependent floral repressor(s). Several studies have reported the epistatic interactions between
Second,
It is noteworthy that CKI and CK2α phosphorylate distinct regions in the PRR37 protein; CKI phosphorylates the recombinant partial proteins PRR37m and PRR37c (Fig. 4B), but CK2α phosphorylates only PRR37m (Fig. 4B). This suggests that the levels of phosphorylation by CKI and CK2α might separately regulate the activity and/or stability of PRR37. Further
Natural variants of
In conclusion, we propose that CKI and CK2α may contribute to enhancing the photoperiod sensitivity of rice through phosphorylation of PRR37. Furthermore, our results provide new, important insights into CKI and/or CK2-mediated phosphorylation of PRR proteins in other plants including barley, wheat, sorghum, maize, and
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