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The genome of the cyanobacterium Synechococcus sp. strain PCC 7942 contains three psbA genes which encode two forms of the D1 protein of photosystem II. Experiments using psbA-lacZ translational fusions and Western blot (immunoblot) analysis have shown that the psbA genes respond differently to changes in light intensity, altering the ratio of the two forms of D1 in the thylakoid membrane. Each gene produces a 1.2-kilobase (kb) mRNA. A probe specific for psbAII transcripts also identified a 1.6-kb mRNA which starts 419 base pairs upstream of the 5' end of the 1.2-kb species and overlaps the entire 1.2-kb transcript. This 419-base-pair region includes an open reading frame (ORF1) of 114 amino acids. We investigated the effects of changes in light intensity on psbAII transcript levels in a series of light shift experiments in the wild-type Synechococcus sp. and in AMC084, a mutant which does not produce the 1.6-kb transcript. After exposure to high light intensities for 15 min, the level of the 1.2-kb psbAII transcript increased in both strains. This transcript was not detected in either strain after transfer to low light intensity. The psbAIII transcript showed the same pattern of response as the 1.2-kb psbAII transcript, whereas the 1.6-kb psbAII transcript was unaffected by different light intensities. The psbAI transcript levels responded oppositely to those of psbAII and psbAIII. These data, considered along with previous results obtained by using lacZ translational gene fusions, indicate that the response of psbA genes to changes in light intensity is controlled primarily at the transcriptional level.

The cyanobacteria Synechococcus sp. strain PCC 7942 and Synechococcus sp. strain PCC 6301 are very closely related and both have been designated by the binomial Anacystis nidulans. The only established difference between the two strains is the superior transformation properties of strain PCC 7942. Significant homology between the rRNA genes of these strains was demonstrated by the ability of an rRNA operon from strain PCC 6301, interrupted by a spectinomycin and streptomycin resistance marker, to transform strain PCC 7942 by recombining with and replacing an endogenous rRNA operon. Restriction fragment length polymorphism data indicated that the chromosomes of the two strains were conserved around the three psbA loci, the two rRNA operons, and the psbDI locus. However, multiple polymorphisms were detected downstream of the psbDII locus, identifying a DNA rearrangement such as an inversion, insertion, or deletion within the chromosome. Analysis of genome structure by pulsed-field gel electrophoresis of large NotI restriction fragments showed only two bands that were visibly shifted between the chromosomes of the two strains. These data support their very close genetic relationship and the feasibility of studying genes derived from strain PCC 6301 in the highly transformable PCC 7942 strain.

The genome of the cyanobacterium Synechococcus sp. PCC7942 contains two genes encoding the D2 polypeptide of photosystem II (PSII), which are designated here as psbDI and psbDII. The psbDI gene, like the psbD gene of plant chloroplasts, is cotranscribed with and overlaps the open reading frame of the psbC gene, encoding the PSII protein CP43. The psbDII gene is not linked to psbC, and appears to be transcribed as a monocistronic message. The two psbD genes encode identical polypeptides of 352 amino acids, which are 86% conserved with the D2 polypeptide of spinach. In plants, the translational start codon of the psbC gene has been reported to be an ATG codon 50 bp upstream from the end of the psbD gene. This triplet is not present in the psbDI sequence of Synechococcus sp., but is replaced by ACG, a codon which is very unlikely to initiate translation. Translation of the psbC gene may begin at a GTG codon which overlap the psbDI open reading frame by 14 bp and is preceded by a block of homology to the 3' end of the 16S ribosomal RNA, a potential ribosome-binding site. There are only two bp differences between the sequences of the two psbD genes; one of these results in substitution in psbDII of GCG for the presumed GTG start codon in psbDI.

It has long been suspected that chloroplasts evolved after an endosymbiotic event involving a photosynthetic prokaryote, presumably a cyanobacterium, and a eukaryotic organism. Recent studies have provided strong evidence about the cyanobacterial nature of chloroplasts. Since the discovery of prochlorophytes, oxygen-evolving photosynthetic prokaryotes containing chlorophyll a and chlorophyll b and lacking phycobiliproteins, there has been speculation that these represent evolutionary intermediates between cyanobacteria and chloroplasts. Prochloron sp., the first described prochlorophyte, proved difficult to work with because it is an obligate symbiont of marine ascidians. Prochlorothrix hollandica, a recently isolated, freshwater filamentous prochlorophyte, is easily maintained in the laboratory. Overall pigment composition and thylakoid membrane structure of P. hollandica suggest it has intermediate characteristics between cyanobacteria and the chloroplasts of higher plants. The P. hollandica psbA genes, which encode the photosystem II thylakoid protein D1, were cloned and sequenced and the sequences compared to those reported for cyanobacteria, a green alga, a liverwort, and several higher plants. The two psbA genes present in P. hollandica encode an identical amino-acid sequence. As in all chloroplast psbA genes, there is a seven amino-acid gap near the C terminus of the derived protein relative to the protein predicted by cyanobacterial genes, suggesting that P. hollandica is part of the lineage that led to chloroplasts after a divergence from cyanobacteria. This hypothesis is also supported by phylogenetic analysis of derived D1 amino-acid sequences from psbA genes of thirteen taxa on the basis of parsimony.

The genome of the cyanobacterium Anacystis nidulans R2 contains three genes (psbA) for the QB protein of photosystem II. This protein is essential for oxygenic photosynthetic electron tansport, and is the target for several herbicides which act by binding directly to the photosynthetic apparatus. Transcripts from the three Anacystis psbA genes are present in wild-type cells at different steady-state levels. The nucleotide sequences of two of the genes, psbAII and psbAIII, predict a protein having the same amino acid sequence which differs from that of the psbAI gene by 25 (out of 360) residues. Inactivation of each of the psbA genes in the Anacystis chromosome, singly or in pairs, shows that each of the genes is capable of producing sufficient functional QB protein to support normal photoautotrophic growth.

The cyanobacterium Synechococcus sp. strain PCC 7942 has two copies of the psbD gene which encodes the D2 polypeptide of the photosystem II (PSII) reaction center. One of the genes, psbDI, overlaps the open reading frame of another photosystem II gene, psbC; the psbDII gene is monocistronic. Gene inactivation experiments had previously shown that psbDII is dispensable under normal laboratory growth conditions. However, similar experiments with psbDI never produced viable psbDI-inactivated mutants, presumably because psbC expression depends on transcription through psbDI. The experiments described here were designed to assess the need for psbDI independent of the need for expression of psbC. A strain, AMC027, was engineered in which a second copy of psbC was expressed from the psbDII locus. Northern (RNA) blot analysis confirmed that both psbDI and psbDII gave rise to dicistronic messages containing psbC sequences in AMC027. In this genetic background, it was possible to inactivate psbDI, creating strain AMC050 and indicating that the psbDII gene is functional. Western immunoblot analysis showed that the products of psbD and psbC, the PSII proteins D2 and CP43, respectively, were present in thylakoids of AMC050, but at reduced levels relative to the wild type, the mutant AMC027, and two psbDII-inactivated mutants. AMC050 consistently formed small colonies on plates and competed poorly in mixed-culture experiments. This suggested that, although not essential for viability, expression from the psbDI locus is required to produce sufficient D2 and CP43 for optimal growth.

Cyanobacteria such as Synechococcus elongatus PCC 7942 exhibit 24-h rhythms of gene expression that are controlled by an endogenous clock that is mechanistically distinct from those described for diverse eukaryotes. Genetic and biochemical experiments over the past decade have identified key components of the circadian oscillator, input pathways that synchronize the clock with the daily environment, and output pathways that relay temporal information to downstream genes. The mechanism of the cyanobacterial circadian clock that is emerging is based principally on the assembly and disassembly of a large complex at whose heart are the proteins KaiA, KaiB, and KaiC. Signal transduction pathways that feed into and out of the clock employ protein domains that are similar to those in two-component regulatory systems of bacteria. [ABSTRACT FROM AUTHOR]