Posttranslational Modification of Maize Chloroplast Pyruvate Orthophosphate Dikinase Reveals the Precise Regulatory Mechanism of Its Enzymatic Activity.
In C4plants, pyruvate orthophosphate dikinase (PPDK) activity is tightly dark/light regulated by reversible phosphorylation of an active-site threonine (Thr) residue; this process is catalyzed by PPDK regulatory protein (PDRP). Phosphorylation and dephosphorylation of PPDK lead to its inactivation and activation, respectively. Here, we show that light intensity rather than the light/dark transition regulates PPDK activity by modulating the reversible phosphorylation at Thr-527 (previously termed Thr-456) of PPDK in maize (Zea mays). The amount of PPDK (unphosphorylated) involved in C4photosynthesis is indeed strictly controlled by light intensity, despite the high levels of PPDK protein that accumulate in mesophyll chloroplasts. In addition, we identified a transit peptide cleavage site, uncovered partial amino-terminal acetylation, and detected phosphorylation at four serine (Ser)/Thr residues, two of which were previously unknown in maize. In vitro experiments indicated that Thr-527 and Ser-528, but not Thr-309 and Ser-506, are targets of PDRP. Modeling suggests that the two hydrogen bonds between the highly conserved residues Ser-528 and glycine-525 are required for PDRP-mediated phosphorylation of the active-site Thr-527 of PPDK. Taken together, our results suggest that the regulation of maize plastid PPDK isoform (C4PPDK) activity is much more complex than previously reported. These diverse regulatory pathways may work alone or in combination to fine-tune C4PPDK activity in response to changes in lighting.
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Xiangshan, Beijing 100093, China (Y.-B.C., J.S., T.-T.B., Q.C., Z.-F.G., Y.-F.W., B.-C.W.);State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China (T.-C.L.);Institute of Basic Medical Sciences, National Center of Biomedical Analysis, Beijing 100850, China (H.-X.W.); andShanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China (X.-G.Z.).
Yi-Bo
YB
Chen
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Xiangshan, Beijing 100093, China (Y.-B.C., J.S., T.-T.B., Q.C., Z.-F.G., Y.-F.W., B.-C.W.);State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China (T.-C.L.);Institute of Basic Medical Sciences, National Center of Biomedical Analysis, Beijing 100850, China (H.-X.W.); andShanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China (X.-G.Z.).
Tian-Cong
TC
Lu
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Xiangshan, Beijing 100093, China (Y.-B.C., J.S., T.-T.B., Q.C., Z.-F.G., Y.-F.W., B.-C.W.);State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China (T.-C.L.);Institute of Basic Medical Sciences, National Center of Biomedical Analysis, Beijing 100850, China (H.-X.W.); andShanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China (X.-G.Z.).
Hong-Xia
HX
Wang
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Xiangshan, Beijing 100093, China (Y.-B.C., J.S., T.-T.B., Q.C., Z.-F.G., Y.-F.W., B.-C.W.);State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China (T.-C.L.);Institute of Basic Medical Sciences, National Center of Biomedical Analysis, Beijing 100850, China (H.-X.W.); andShanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China (X.-G.Z.).
Jie
J
Shen
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Xiangshan, Beijing 100093, China (Y.-B.C., J.S., T.-T.B., Q.C., Z.-F.G., Y.-F.W., B.-C.W.);State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China (T.-C.L.);Institute of Basic Medical Sciences, National Center of Biomedical Analysis, Beijing 100850, China (H.-X.W.); andShanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China (X.-G.Z.).
Tian-Tian
TT
Bu
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Xiangshan, Beijing 100093, China (Y.-B.C., J.S., T.-T.B., Q.C., Z.-F.G., Y.-F.W., B.-C.W.);State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China (T.-C.L.);Institute of Basic Medical Sciences, National Center of Biomedical Analysis, Beijing 100850, China (H.-X.W.); andShanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China (X.-G.Z.).
Qing
Q
Chao
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Xiangshan, Beijing 100093, China (Y.-B.C., J.S., T.-T.B., Q.C., Z.-F.G., Y.-F.W., B.-C.W.);State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China (T.-C.L.);Institute of Basic Medical Sciences, National Center of Biomedical Analysis, Beijing 100850, China (H.-X.W.); andShanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China (X.-G.Z.).
Zhi-Fang
ZF
Gao
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Xiangshan, Beijing 100093, China (Y.-B.C., J.S., T.-T.B., Q.C., Z.-F.G., Y.-F.W., B.-C.W.);State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China (T.-C.L.);Institute of Basic Medical Sciences, National Center of Biomedical Analysis, Beijing 100850, China (H.-X.W.); andShanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China (X.-G.Z.).
Xin-Guang
XG
Zhu
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Xiangshan, Beijing 100093, China (Y.-B.C., J.S., T.-T.B., Q.C., Z.-F.G., Y.-F.W., B.-C.W.);State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China (T.-C.L.);Institute of Basic Medical Sciences, National Center of Biomedical Analysis, Beijing 100850, China (H.-X.W.); andShanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China (X.-G.Z.).
Yue-Feng
YF
Wang
Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Xiangshan, Beijing 100093, China (Y.-B.C., J.S., T.-T.B., Q.C., Z.-F.G., Y.-F.W., B.-C.W.);State Key Laboratory of Plant Genomics and National Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China (T.-C.L.);Institute of Basic Medical Sciences, National Center of Biomedical Analysis, Beijing 100850, China (H.-X.W.); andShanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China (X.-G.Z.) wangbc@ibcas.ac.cn.
Bai-Chen
http://orcid.org/0000-0003-0169-4393
BC
Wang