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Identification of factors involved in the biogenesis of thylakoid membranes in cyanobacteria
Identification of factors involved in the biogenesis of thylakoid membranes in cyanobacteria
Photosynthesis is the key reaction that has enabled the evolution of complex life on earth. The gram-negative cyanobacterium Synechocystis sp. PCC 6803 (hereinafter Synechocystis) is a well-established model organism for research on photosynthesis. The Synechocystis cell is surrounded by an outer membrane and a plasma membrane. Photosynthesis takes place in the thylakoid membranes (TMs) localised in the cytoplasm, which are shaped towards the plasma membrane and form the converging zone. The resulting converging zone is the region of early Photosystem II (PSII) assembly steps. The precursor form of D1 (pD1) is preloaded with manganese by the processing associated tetracopeptide (TPR) protein (PratA) in a subcompartment of the TM. The PratA defined membrane (PDM) represents the converging zones. Moreover, PratA forms a 200 kDa complex with the high temperature requirement homologue A (HhoA) and pilin protein Q (PilQ) in the periplasm. Beside PratA, the protein Curvature Thylakoid1 (CurT) plays an important role in the formation of the converging zones. A reduced photosynthetic performance and the absence of convergence zones is caused by knocking out CurT. This photosynthetic phenotype can be suppressed by the formation of suppressor lines of curT–: sucurT-. To get a better understanding of the convergence zones two different screening methods were performed: a proteomic screen with the focus on PratA and a genetic screen focused on curT- and its suppressor sucurT-. In the primary analysis some selected candidates of the screen showed defects in photosynthetic performance and growth. The open reading frame (ORF) slr2070 encodes for a protein named anchoring of convergence membranes (AncM). AncM was found independent in both screens. Concerning this, AncM is reduced in pratA- and co-expressed with CurT. In sucurT- ancM contains a STOP codon in the N-terminus at the amino acid position 164 in front of a transmembrane domain (TMD). Based on that, AncM was not detected in sucurT- by immunoblotting. Immunofluorescence and sucrose density gradients localised the membrane protein AncM at the PDM. Moreover a photosynthetic phenotype was observed in ancM-. In some regions AncM is colocalised with CurT. Electron microscopy studies of ancM- shows thylakoids that are not connected to the plasma membrane. This reveals a structuring function of AncM relating to CurT. It is suggested that AncM anchors the thylakoids and offers a fixing point for the thylakoids at the converging zones. Based on the important function of AncM at the converging zones the independent screens offer a powerful method to investigate convergence zones as a specialised compartment of the thylakoids.
Photosynthesis, Thylakoids, Cynanobacteria
Hamm, Julia
2021
English
Universitätsbibliothek der Ludwig-Maximilians-Universität München
Hamm, Julia (2021): Identification of factors involved in the biogenesis of thylakoid membranes in cyanobacteria. Dissertation, LMU München: Faculty of Biology
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Abstract

Photosynthesis is the key reaction that has enabled the evolution of complex life on earth. The gram-negative cyanobacterium Synechocystis sp. PCC 6803 (hereinafter Synechocystis) is a well-established model organism for research on photosynthesis. The Synechocystis cell is surrounded by an outer membrane and a plasma membrane. Photosynthesis takes place in the thylakoid membranes (TMs) localised in the cytoplasm, which are shaped towards the plasma membrane and form the converging zone. The resulting converging zone is the region of early Photosystem II (PSII) assembly steps. The precursor form of D1 (pD1) is preloaded with manganese by the processing associated tetracopeptide (TPR) protein (PratA) in a subcompartment of the TM. The PratA defined membrane (PDM) represents the converging zones. Moreover, PratA forms a 200 kDa complex with the high temperature requirement homologue A (HhoA) and pilin protein Q (PilQ) in the periplasm. Beside PratA, the protein Curvature Thylakoid1 (CurT) plays an important role in the formation of the converging zones. A reduced photosynthetic performance and the absence of convergence zones is caused by knocking out CurT. This photosynthetic phenotype can be suppressed by the formation of suppressor lines of curT–: sucurT-. To get a better understanding of the convergence zones two different screening methods were performed: a proteomic screen with the focus on PratA and a genetic screen focused on curT- and its suppressor sucurT-. In the primary analysis some selected candidates of the screen showed defects in photosynthetic performance and growth. The open reading frame (ORF) slr2070 encodes for a protein named anchoring of convergence membranes (AncM). AncM was found independent in both screens. Concerning this, AncM is reduced in pratA- and co-expressed with CurT. In sucurT- ancM contains a STOP codon in the N-terminus at the amino acid position 164 in front of a transmembrane domain (TMD). Based on that, AncM was not detected in sucurT- by immunoblotting. Immunofluorescence and sucrose density gradients localised the membrane protein AncM at the PDM. Moreover a photosynthetic phenotype was observed in ancM-. In some regions AncM is colocalised with CurT. Electron microscopy studies of ancM- shows thylakoids that are not connected to the plasma membrane. This reveals a structuring function of AncM relating to CurT. It is suggested that AncM anchors the thylakoids and offers a fixing point for the thylakoids at the converging zones. Based on the important function of AncM at the converging zones the independent screens offer a powerful method to investigate convergence zones as a specialised compartment of the thylakoids.