The chloroplast calcium sensor protein CAS is part of the STN7/STN8 kinase phosphorylation network and is required for photoacclimation
The chloroplast calcium sensor protein CAS is part of the STN7/STN8 kinase phosphorylation network and is required for photoacclimation
View/Type of Scholarly Publication
DissertationDate of Exam
21.03.2019Date of Publication
10.03.2020Advisor
Vothknecht, UteCo-Referee
Bartels, DorotheaMetadata
Show full item record
Citable Links 
Abstract
In the present work, the phosphorylation profile of the chloroplast-localized Calcium Sensing Receptor (CAS) from A. thaliana was investigated and it could be shown that CAS is a target of multiple protein kinases acting differentially on several residues. Phosphoproteomics followed up by biochemical kinase assays strongly indicated that CAS is part of a phosphorylation network involving the important state transistion kinases STN7 and STN8 as well as, at least, one other calcium-regulated protein kinase. The role of light and calcium behind the activation of CAS phosphorylation was investigated in vitro in more detail and revealed that, at least under normal growth light conditions, STN7 might be the major kinase acting on CAS.
The analysis was extended at the level of individual residues by studying the phylogenetic conservation of experimentally described phosphoresidues of the A. thaliana CAS isoform and by conducting in vitro assays using recombinant CAS fragments carrying mutations at selected positions. These analyses confirmed the relevance of the previously described phosphorylation site Thr-380, but also showed that other residues, in particular Thr-376, are possible targets.
A spectrometric analysis of cas mutant plants revealed that CAS is very likely involved in the processes of photoacclimation to high light, as evidenced by a persistent strong excitation of PSI under this condition. The analysis of the phosphorylation status of several known thylakoid phosphoproteins in the cas mutant further suggested a potential defect in the dephosphorylation of the important light harvesting protein LHCII under high irradiance, suggesting a possible role for CAS in mediating the activity of the TAP38 phosphatase.
In addition to the post-translational modification of CAS, a potential involvement of CAS in the circadian network of the chloroplast was explored. RT-PCR analyses revealed that the transcription of the CAS follows a regular diurnal rhythm, with the highest levels of its transcript levels found at the end of the night and lowest levels at the end of the day. Interestingly, the levels of the CAS protein appear to follow an opposite, 12 hours shifted rhythm, suggesting a physiological requirement for higher abundance of CAS during the day. In light of these results, a possible working model is discussed that integrates the evidences on the phosphorylation profiles and the diurnal regulation with a suggested involvement of CAS in photoacclimation responses.
The analysis was extended at the level of individual residues by studying the phylogenetic conservation of experimentally described phosphoresidues of the A. thaliana CAS isoform and by conducting in vitro assays using recombinant CAS fragments carrying mutations at selected positions. These analyses confirmed the relevance of the previously described phosphorylation site Thr-380, but also showed that other residues, in particular Thr-376, are possible targets.
A spectrometric analysis of cas mutant plants revealed that CAS is very likely involved in the processes of photoacclimation to high light, as evidenced by a persistent strong excitation of PSI under this condition. The analysis of the phosphorylation status of several known thylakoid phosphoproteins in the cas mutant further suggested a potential defect in the dephosphorylation of the important light harvesting protein LHCII under high irradiance, suggesting a possible role for CAS in mediating the activity of the TAP38 phosphatase.
In addition to the post-translational modification of CAS, a potential involvement of CAS in the circadian network of the chloroplast was explored. RT-PCR analyses revealed that the transcription of the CAS follows a regular diurnal rhythm, with the highest levels of its transcript levels found at the end of the night and lowest levels at the end of the day. Interestingly, the levels of the CAS protein appear to follow an opposite, 12 hours shifted rhythm, suggesting a physiological requirement for higher abundance of CAS during the day. In light of these results, a possible working model is discussed that integrates the evidences on the phosphorylation profiles and the diurnal regulation with a suggested involvement of CAS in photoacclimation responses.
Classification (DDC)
580 Pflanzen (Botanik)Cutolo, Edoardo Andrea: The chloroplast calcium sensor protein CAS is part of the STN7/STN8 kinase phosphorylation network and is required for photoacclimation. - Bonn, 2020. - Dissertation, Rheinische Friedrich-Wilhelms-Universität Bonn.
Online-Ausgabe in bonndoc: https://nbn-resolving.org/urn:nbn:de:hbz:5-54373
Online-Ausgabe in bonndoc: https://nbn-resolving.org/urn:nbn:de:hbz:5-54373
@phdthesis{handle:20.500.11811/8244,
urn: https://nbn-resolving.org/urn:nbn:de:hbz:5-54373,
author = {{Edoardo Andrea Cutolo}},
title = {The chloroplast calcium sensor protein CAS is part of the STN7/STN8 kinase phosphorylation network and is required for photoacclimation},
school = {Rheinische Friedrich-Wilhelms-Universität Bonn},
year = 2020,
month = mar,
note = {In the present work, the phosphorylation profile of the chloroplast-localized Calcium Sensing Receptor (CAS) from A. thaliana was investigated and it could be shown that CAS is a target of multiple protein kinases acting differentially on several residues. Phosphoproteomics followed up by biochemical kinase assays strongly indicated that CAS is part of a phosphorylation network involving the important state transistion kinases STN7 and STN8 as well as, at least, one other calcium-regulated protein kinase. The role of light and calcium behind the activation of CAS phosphorylation was investigated in vitro in more detail and revealed that, at least under normal growth light conditions, STN7 might be the major kinase acting on CAS.
The analysis was extended at the level of individual residues by studying the phylogenetic conservation of experimentally described phosphoresidues of the A. thaliana CAS isoform and by conducting in vitro assays using recombinant CAS fragments carrying mutations at selected positions. These analyses confirmed the relevance of the previously described phosphorylation site Thr-380, but also showed that other residues, in particular Thr-376, are possible targets.
A spectrometric analysis of cas mutant plants revealed that CAS is very likely involved in the processes of photoacclimation to high light, as evidenced by a persistent strong excitation of PSI under this condition. The analysis of the phosphorylation status of several known thylakoid phosphoproteins in the cas mutant further suggested a potential defect in the dephosphorylation of the important light harvesting protein LHCII under high irradiance, suggesting a possible role for CAS in mediating the activity of the TAP38 phosphatase.
In addition to the post-translational modification of CAS, a potential involvement of CAS in the circadian network of the chloroplast was explored. RT-PCR analyses revealed that the transcription of the CAS follows a regular diurnal rhythm, with the highest levels of its transcript levels found at the end of the night and lowest levels at the end of the day. Interestingly, the levels of the CAS protein appear to follow an opposite, 12 hours shifted rhythm, suggesting a physiological requirement for higher abundance of CAS during the day. In light of these results, a possible working model is discussed that integrates the evidences on the phosphorylation profiles and the diurnal regulation with a suggested involvement of CAS in photoacclimation responses.},
url = {https://hdl.handle.net/20.500.11811/8244}
}
urn: https://nbn-resolving.org/urn:nbn:de:hbz:5-54373,
author = {{Edoardo Andrea Cutolo}},
title = {The chloroplast calcium sensor protein CAS is part of the STN7/STN8 kinase phosphorylation network and is required for photoacclimation},
school = {Rheinische Friedrich-Wilhelms-Universität Bonn},
year = 2020,
month = mar,
note = {In the present work, the phosphorylation profile of the chloroplast-localized Calcium Sensing Receptor (CAS) from A. thaliana was investigated and it could be shown that CAS is a target of multiple protein kinases acting differentially on several residues. Phosphoproteomics followed up by biochemical kinase assays strongly indicated that CAS is part of a phosphorylation network involving the important state transistion kinases STN7 and STN8 as well as, at least, one other calcium-regulated protein kinase. The role of light and calcium behind the activation of CAS phosphorylation was investigated in vitro in more detail and revealed that, at least under normal growth light conditions, STN7 might be the major kinase acting on CAS.
The analysis was extended at the level of individual residues by studying the phylogenetic conservation of experimentally described phosphoresidues of the A. thaliana CAS isoform and by conducting in vitro assays using recombinant CAS fragments carrying mutations at selected positions. These analyses confirmed the relevance of the previously described phosphorylation site Thr-380, but also showed that other residues, in particular Thr-376, are possible targets.
A spectrometric analysis of cas mutant plants revealed that CAS is very likely involved in the processes of photoacclimation to high light, as evidenced by a persistent strong excitation of PSI under this condition. The analysis of the phosphorylation status of several known thylakoid phosphoproteins in the cas mutant further suggested a potential defect in the dephosphorylation of the important light harvesting protein LHCII under high irradiance, suggesting a possible role for CAS in mediating the activity of the TAP38 phosphatase.
In addition to the post-translational modification of CAS, a potential involvement of CAS in the circadian network of the chloroplast was explored. RT-PCR analyses revealed that the transcription of the CAS follows a regular diurnal rhythm, with the highest levels of its transcript levels found at the end of the night and lowest levels at the end of the day. Interestingly, the levels of the CAS protein appear to follow an opposite, 12 hours shifted rhythm, suggesting a physiological requirement for higher abundance of CAS during the day. In light of these results, a possible working model is discussed that integrates the evidences on the phosphorylation profiles and the diurnal regulation with a suggested involvement of CAS in photoacclimation responses.},
url = {https://hdl.handle.net/20.500.11811/8244}
}
The following license files are associated with this item:
