Abstract
Deletion of the genes for four or five small Cab-like proteins (SCPs) in photosystem (PS) I-less and PS I-less/PS II-less strains of Synechocystis sp. PCC 6803 caused a large decrease in the chlorophyll and carotenoid content of the cells without accumulation of early intermediates in the chlorophyll biosynthesis pathway, suggesting limited chlorophyll availability. The PS II/PS I ratio increased upon deletion of multiple SCPs in a wild type background, similar to what is observed in the presence of subsaturating concentrations of gabaculin, an inhibitor of an early step in the tetrapyrrole biosynthesis pathway. Upon deletion of multiple SCPs, neither 77 K fluorescence emission properties of phycobilisomeless thylakoids from the PS I-less/PS II-less strain nor the energy trapping efficiency of PS II were affected, indicating that under steady-state conditions SCPs do not bind much chlorophyll and do not serve as PS II antenna. Under conditions where protochlorophyllide reduction and thus chlorophyll synthesis were inhibited, chlorophyll disappeared quickly in a mutant lacking all five SCPs. This implies a role of SCPs in stabilization of chlorophyll-binding proteins and/or in reuse of chlorophylls. Under these conditions of inhibited reduction of protochlorophyllide, the accumulation kinetics of this intermediate were greatly altered in the absence of the five SCPs. This indicates an alteration of tetrapyrrole biosynthesis kinetics by SCPs. Based on this and other evidence, we propose that SCPs bind carotenoids and transiently bind chlorophyll, aiding in the supply of chlorophyll to nascent or reassembling photosynthetic complexes, and regulate the tetrapyrrole biosynthesis pathway as a function of the demand for chlorophyll.
Highlights
In a previous paper devoted to the function of SCPs in Synechocystis, we hypothesized that SCPs may be involved in regulation of the tetrapyrrole biosynthesis pathway depending on the occupancy of their pigment binding sites [13]
This interpretation was based on the observations that PChlide and phycobilin levels decreased in the single ScpB or ScpE deletion mutants grown under LAHG conditions, and recovery of chlorophyll and phycobilin levels was slowed down in the deletion mutants compared with the control strain after the cultures were transferred to continuous light
Slow pigment synthesis upon transfer of LAHG-grown PS I-less/chlLϪ cells to continuous light was found in the PS I-less/chlLϪ mutant that lacked both scpC and scpD (Fig. 6A) but not when either scpC or scpD was missing,2 suggesting that these two proteins, which share about 87% identity in amino acid sequence, have a complementary regulatory function in chlorophyll synthesis
Summary
PCC 6803 scpB coding region and 46 bp downstream of the stop codon) was deleted from the plasmid containing the scpB gene with 440 and 820 bp upstream and downstream, respectively, of this gene [13] and was replaced by a zeocin resistance cassette [16]. Transformants were selected by screening for resistance to appropriate antibiotics and subcultured at increasing concentrations of antibiotics to allow segregation of wild-type and mutant genome copies to occur, leading to homozygous strains in wild-type, PS I-less, PS II-less (psbBϪ/psbCϪ), and PS I-less/PS II-less backgrounds that are unable to make four or all five SCPs. To remove the erythromycin resistance-sacB cassette from the genomes of scpC interruption mutants, homozygous mutant cells were transformed with the markerless scpC deletion construct. A small amount of phycobilins remained in the preparations, as seen from 77 K fluorescence emission and excitation spectra
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