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Mol. Cells 2013; 35(5): 381-387

Published online May 31, 2013

https://doi.org/10.1007/s10059-013-2185-0

© The Korean Society for Molecular and Cellular Biology

Rice Small C2-Domain Proteins Are Phosphorylated by Calcium-Dependent Protein Kinase

Chang Ho Kang, Byeong Cheol Moon, Hyeong Cheol Park, Sung Cheol Koo, Yong Hun Chi, Yong Hwa Cheong, Byung-Dae Yoon, Sang Yeol Lee, and Cha Young Kim

Bio-Materials Research Institute, Korea Research Institute of Bioscience and Biotechnology, Jeongeup 580-185, Korea, 1Division of Applied Life Sciences (Brain Korea 21 Program), Plant Molecular Biology and Biotechnology Research Center, Gyeongsang National University, Jinju 660-701, Korea, 2Center of Herbal Resources Research, Korea Institute of Oriental Medicine, Daejeon 305-811, Korea, 3Department of Bio-Environmental Science, Sunchon National University, Suncheon 540-742, Korea, 4Department of Functional Crop, National Institute of Crop Science, Rural Development Administration, Miryang 627-803, Korea

Received: July 20, 2013; Revised: January 21, 2013; Accepted: January 22, 2013

Abstract

We previously reported that OsERG1 and OsERG3 encode rice small C2-domain proteins with different biochemical properties in Ca(2+)- and phospholipid-binding assays. Os-ERG1 exhibited Ca(2+)-dependent phospholipid binding, which was not observed with OsERG3. In the present study, we show that both OsERG1 and OsERG3 proteins exhibit oligomerization properties as determined by native polyacrylamide gel electrophoresis (PAGE) and glutaraldehyde cross-linking experiments. Furthermore, in vitro phosphorylation assays reveal the phosphorylation of OsERG1 and OsERG3 by a rice calcium-dependent protein kinase, OsCDPK5. Our mutation analysis on putative serine phosphorylation sites shows that the first serine (Ser) at position 41 of OsERG1 may be an essential residue for phosphorylation by OsCDPK5. Mutation of Ser41 to alanine (OsERG1S41A) and aspartate (OsERG1S41D) abolishes the ability of OsERG1 to bind phospholipids regardless of the presence or absence of Ca(2+) ions. In addition, unlike the OsERG1 wild-type form, the mutant OsERG1 (S41A)::smGFP construct lost the ability to translocate from the cytosol to the plasma membrane in response to calcium ions or fungal elicitor. These results indicate that Ser41 may be essential for the function of OsERG1.

Keywords Ca(2+)/phospholipid-binding, calcium-dependent protein kinase, oligomerization, phosphorylation, small C2-domain protein

Article

Research Article

Mol. Cells 2013; 35(5): 381-387

Published online May 31, 2013 https://doi.org/10.1007/s10059-013-2185-0

Copyright © The Korean Society for Molecular and Cellular Biology.

Rice Small C2-Domain Proteins Are Phosphorylated by Calcium-Dependent Protein Kinase

Chang Ho Kang, Byeong Cheol Moon, Hyeong Cheol Park, Sung Cheol Koo, Yong Hun Chi, Yong Hwa Cheong, Byung-Dae Yoon, Sang Yeol Lee, and Cha Young Kim

Bio-Materials Research Institute, Korea Research Institute of Bioscience and Biotechnology, Jeongeup 580-185, Korea, 1Division of Applied Life Sciences (Brain Korea 21 Program), Plant Molecular Biology and Biotechnology Research Center, Gyeongsang National University, Jinju 660-701, Korea, 2Center of Herbal Resources Research, Korea Institute of Oriental Medicine, Daejeon 305-811, Korea, 3Department of Bio-Environmental Science, Sunchon National University, Suncheon 540-742, Korea, 4Department of Functional Crop, National Institute of Crop Science, Rural Development Administration, Miryang 627-803, Korea

Received: July 20, 2013; Revised: January 21, 2013; Accepted: January 22, 2013

Abstract

We previously reported that OsERG1 and OsERG3 encode rice small C2-domain proteins with different biochemical properties in Ca(2+)- and phospholipid-binding assays. Os-ERG1 exhibited Ca(2+)-dependent phospholipid binding, which was not observed with OsERG3. In the present study, we show that both OsERG1 and OsERG3 proteins exhibit oligomerization properties as determined by native polyacrylamide gel electrophoresis (PAGE) and glutaraldehyde cross-linking experiments. Furthermore, in vitro phosphorylation assays reveal the phosphorylation of OsERG1 and OsERG3 by a rice calcium-dependent protein kinase, OsCDPK5. Our mutation analysis on putative serine phosphorylation sites shows that the first serine (Ser) at position 41 of OsERG1 may be an essential residue for phosphorylation by OsCDPK5. Mutation of Ser41 to alanine (OsERG1S41A) and aspartate (OsERG1S41D) abolishes the ability of OsERG1 to bind phospholipids regardless of the presence or absence of Ca(2+) ions. In addition, unlike the OsERG1 wild-type form, the mutant OsERG1 (S41A)::smGFP construct lost the ability to translocate from the cytosol to the plasma membrane in response to calcium ions or fungal elicitor. These results indicate that Ser41 may be essential for the function of OsERG1.

Keywords: Ca(2+)/phospholipid-binding, calcium-dependent protein kinase, oligomerization, phosphorylation, small C2-domain protein

Mol. Cells
Nov 30, 2022 Vol.45 No.11, pp. 763~867
COVER PICTURE
Naive (cyan) and axotomized (magenta) retinal ganglion cell axons in Xenopus tropicalis (Choi et al., pp. 846-854).

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