Eukaryotic-like Ser/Thr Protein Kinases SpkC/F/K Are Involved in Phosphorylation of GroES in the Cyanobacterium Synechocystis

Abstract

Serine/threonine protein kinases (STPKs) are the major participants in intracellular signal transduction in eukaryotes, such as yeasts, fungi, plants, and animals. Genome sequences indicate that these kinases are also present in prokaryotes, such as cyanobacteria. However, their roles in signal transduction in prokaryotes remain poorly understood. We have attempted to identify the roles of STPKs in response to heat stress in the prokaryotic cyanobacterium Synechocystis sp. PCC 6803, which has 12 genes for STPKs. Each gene was individually inactivated to generate a gene-knockout library of STPKs. We applied in vitro Ser/Thr protein phosphorylation and phosphoproteomics and identified the methionyl-tRNA synthetase, large subunit of RuBisCO, 6-phosphogluconate dehydrogenase, translation elongation factor Tu, heat-shock protein GrpE, and small chaperonin GroES as the putative targets for Ser/Thr phosphorylation. The expressed and purified GroES was used as an external substrate to screen the protein extracts of the individual mutants for their Ser/Thr kinase activities. The mutants that lack one of the three protein kinases, SpkC, SpkF, and SpkK, were unable to phosphorylate GroES in vitro, suggesting possible interactions between them towards their substrate. Complementation of the mutated SpkC, SpkF, and SpkK leads to the restoration of the ability of cells to phosphorylate the GroES. This suggests that these three STPKs are organized in a sequential order or a cascade and they work one after another to finally phosphorylate the GroES.

Interplay of SpkG kinase and the Slr0151 protein in the phosphorylation of ferredoxin 5 in Synechocystis sp. strain PCC 6803

“…In the genome of the model unicellular cyanobacterium Synechocystis sp. PCC6803 (hereafter Synechocystis 6803), seven genes, spkA–spkG , have been predicted to encode for the S/T protein kinases of the Pkn2 family, and five genes, spkH–spkL , for the ones of the ABC1 family . SpkA is involved in the control of cell motility by regulating expression of pili family genes .…”

“…Zorina et al . demonstrated that SpkC, SpkF and SpkK are involved in phosphorylation of the small chaperonin GroES. However, regulation, function, and substrate recognition of Synechocystis 6803 S/T protein kinases remain poorly investigated .…”

“…SpkE is probably involved in the regulation of nitrogen metabolism [18], while SpkG was shown to play role in salt stress response, positively regulating expression of 15 genes [19]. Zorina et al [14] demonstrated that SpkC, SpkF and SpkK are involved in phosphorylation of the small chaperonin GroES. However, regulation, function, and substrate recognition of Synechocystis 6803 S/T protein kinases remain poorly investigated [13].…”

Thiol-Based Redox Modulation of a Cyanobacterial Eukaryotic-Type Serine/Threonine Kinase Required for Oxidative Stress Tolerance

“…However, SpkE lacks several key amino acids that are necessary for ATP binding and is not a true protein kinase (15). In addition, there are five putative kinases of the ABC1-type, named SpkH through L (50). The functions of a few cyanobacterial Ser/Thr kinases have been addressed through phenotypical characterization of the respective mutants.…”

Transcriptomic analysis of Synechocystis sp. PCC6803 under low-temperature stress

“…SpkG can sense high-salt stress signals directly (Liang et al, 2011), and spkG was found to be highly down-regulated in iron defi ciency under peroxide stress (Singh et al, 2004). In addition, SpkC, SpkF, and SpkK work one after the other to fi nally phosphorylate the co-chaperonin GroES (Zorina et al, 2011).…”