Twitching motility is a flagella-independent form of bacterial translocation over moist surfaces. It occurs by the extension, tethering, and then retraction of polar type IV pili, which operate in a manner similar to a grappling hook. Twitching motility is equivalent to social gliding motility in Myxococcus xanthus and is important in host colonization by a wide range of plant and animal pathogens, as well as in the formation of biofilms and fruiting bodies. The biogenesis and function of type IV pili is controlled by a large number of genes, almost 40 of which have been identified in Pseudomonas aeruginosa. A number of genes required for pili assembly are homologous to genes involved in type II protein secretion and competence for DNA uptake, suggesting that these systems share a common architecture. Twitching motility is also controlled by a range of signal transduction systems, including two-component sensor-regulators and a complex chemosensory system.

Type IV pili and twitching motility.

Naturally competent bacteria are able to take up exogenous DNA and undergo genetic transformation. The transport of DNA from the extracellular milieu into the cytoplasm is a complex process, and requires proteins that are related to those involved in the assembly of type IV pili and type II secretion systems, as well as a DNA translocase complex at the cytoplasmic membrane. Here, we will review the current knowledge of DNA transport during transformation.

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DNA Uptake During Bacterial Transformation

DNA microarrays bearing nearly all of the genes of the unicellular cyanobacterium Synechocystis sp PCC 6803 were used to examine the temporal program of gene expression during acclimation from low to high light intensity. A complete pattern is provided of gene expression during acclimation of a photosynthetic organism to changing light intensity. More than 160 responsive genes were identified and classified into distinct sets. Genes involved in light absorption and photochemical reactions were downregulated within 15 min of exposure to high light intensity, whereas those associated with CO2 fixation and protection from photoinhibition were upregulated. Changes in the expression of genes involved in replication, transcription, and translation, which were induced to support cellular proliferation, occurred later. Several unidentified open reading frames were induced or repressed. The possible involvement of these genes in the acclimation to high light conditions is discussed.

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DNA Microarray Analysis of Cyanobacterial Gene Expression during Acclimation to High Light

Cyanobacteria may possess two distinct nickel-iron (NiFe)-hydrogenases: an uptake enzyme found in N(2)-fixing strains, and a bidirectional one present in both non-N(2)-fixing and N(2)-fixing strains. The uptake hydrogenase (encoded by hupSL) catalyzes the consumption of the H(2) produced during N(2) fixation, while the bidirectional enzyme (hoxEFUYH) probably plays a role in fermentation and/or acts as an electron valve during photosynthesis. hupSL constitute a transcriptional unit, and are essentially transcribed under N(2)-fixing conditions. The bidirectional hydrogenase consists of a hydrogenase and a diaphorase part, and the corresponding five hox genes are not always clustered or cotranscribed. The biosynthesis/maturation of NiFe-hydrogenases is highly complex, requiring several core proteins. In cyanobacteria, the genes that are thought to affect hydrogenases pleiotropically (hyp), as well as the genes presumably encoding the hydrogenase-specific endopeptidases (hupW and hoxW) have been identified and characterized. Furthermore, NtcA and LexA have been implicated in the transcriptional regulation of the uptake and the bidirectional enzyme respectively. Recently, the phylogenetic origin of cyanobacterial and algal hydrogenases was analyzed, and it was proposed that the current distribution in cyanobacteria reflects a differential loss of genes according to their ecological needs or constraints. In addition, the possibilities and challenges of cyanobacterial-based H(2) production are addressed.

Cyanobacterial hydrogenases: diversity, regulation and applications

A new group of photoreceptors has been experimentally revealed in cyanobacteria. They are phototaxis regulator SyPixJ1, TePixJ and AnPixJ, chromatic acclimation regulator SyCcaS, circadian input kinase homolog SyCikA and many other candidates, which have been found only in cyanobacteria to date. These new photoreceptors are now proposed to be “cyanobacteriochromes”. They are characterized by the presence of a chromophore-binding GAF domain that is homologous to the tetrapyrrole-binding GAF domain of the phytochrome. Here, we summarized unique features of those representatives: (1) only the GAF domain is sufficient for full photoconversion, (2) the GAF domain is homologous to but distinct from the phytochrome GAF, (3) the GAF domain binds a linear tetrapyrrole pigment such as phycoviolobilin or phycocyanobilin, (4) spectral properties are very diverse from near ultra-violet to red region. We also discussed the functionality of the other candidate GAFs, structure and evolution.

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Cyanobacteriochromes: a new superfamily of tetrapyrrole-binding photoreceptors in cyanobacteria

;The relevance of pilus-related genes to motility, pilus structure on the cell surface and competency of natural transformation was studied by gene disruption analysis in the unicellular motile cyanobacterium Synechocystis sp. PCC 6803. The genes disrupted in this study were chosen as related to the pil genes for biogenesis of the type IV pili in a Gram-negative bacterium Pseudomonas aeruginosa .I t was found that motility of Synechocystis cells was lost in the mutants of slr0063, slr1274, slr1275, slr1276, slr1277 and sll1694 together with a simultaneous loss of the thick pili on the cell surface. Competency of the natural transformation was lost in the mutants listed above and slr0197-disruptant. The gene slr0197 was previously predicted as a competence gene by a search with sequence-independent DNAbinding structure [Yura et al. (1999) DNA Res. 6: 75]. It was suggested that both DNA uptake for natural transformation and motility are mediated by a specific type IV-like pilus structure, while a putative DNA-binding protein encoded by slr0197 is additionally required for the DNA uptake. Based on the homology with the pil genes in P. aeruginosa, slr0063, slr1274, slr1275, slr1276, slr1277 and sll1694 were designated pilB1, pilM, pilN, pilO, pilQ and pilA1, respectively. The gene slr0197 was designated comA.

Mutational analysis of genes involved in pilus structure, motility and transformation competency in the unicellular motile cyanobacterium Synechocystis sp. PCC 6803.

Synechocystis: sp. PCC 6803 is a unicellular motile cyanobacterium, which shows positive or negative phototaxis on agar plates under lateral illumination. By gene disruption in a substrain showing of positive phototaxis, it was demonstrated that mutants defective in sll0038, sll0039, sll0041, sll0042 or sll0043 lost positive phototaxis but showed negative phototaxis away from the light source. Mutants of sll0040, which is located within the cluster of these genes, retained the capacity of positive phototaxis but to a lesser extent than the parent cells. These genes are homologous to che genes, which are involved in flagellar switching for bacterial chemotaxis. Interestingly, sll0041 (designated pisJ1) is predicted to have a chromophore-binding motif of phytochrome-like proteins and a signaling motif of chemoreceptors for bacterial chemotaxis. It is strongly suggested that the positive phototactic response was mediated by a phytochrome-like photoreceptor and CheA/CheY-type signal transduction system.

Novel Putative Photoreceptor and Regulatory Genes Required for the Positive Phototactic Movement of the Unicellular Motile Cyanobacterium Synechocystis sp. PCC 6803

The genome of the unicellular cyanobacterium Synechocystis sp. strain PCC 6803 comprises many open reading frames (ORFs) which putatively encode eukaryotic-type protein kinase and protein phosphatase. Based on gene disruption analysis, a region of the hypothetical ORF sll1575, which retained a part of the protein kinase motif, was found to be required for normal motility in the original isolate of strain PCC 6803. Sequence determination revealed that in this strain sll1575 was part of a gene (designated spkA) which harbored an entire eukaryotic-type Ser/Thr protein kinase motif. Strain ATCC 27184 and a glucose-tolerant strain derived from the same isolate as the PCC strain had a frameshift mutation dividing spkA into ORFs sll1574 and sll1575. The structural integrity of spkA agreed well with the motility phenotype, determined by colony morphology on agar plates. The spkA gene was expressed in Escherichia coli as a His-tagged protein, which was purified by Ni 21 affinity chromatography. With [g- 32 P]ATP, SpkA was autophosphorylated and transferred the phosphate group to casein, myelin basic protein, and histone. SpkA also phosphorylated several proteins in the membrane fraction of Synechocystis cells. These results suggest that SpkA is a eukaryotic-type Ser/Thr protein kinase and regulates cellular motility via phosphorylation of the membrane proteins in Synechocystis. Protein phosphorylation-dephosphorylation is a mechanism widely used to regulate proteins. In prokaryotes, phosphotransfer of the protein His kinase to the Asp residue in the

A Eukaryotic-Type Protein Kinase, SpkA, Is Required for Normal Motility of the Unicellular Cyanobacterium Synechocystis sp. Strain PCC 6803

Structural role of the second copy of the rod–core linker CpcG, which was found by genome analysis, was studied in Synechocystis sp. PCC 6803 by gene disruption and fractionation of phycobilisome (sub)complexes. Disruption of cpcG2 (sll1471) resulted in a marked decrease in phycocyanin content both in the background of wild-type and cpcG1 (slr2051)-disruptant. The unique phycocyanin rod–CpcG2 complex without the major allophycocyanin components was isolated from the cpcG1-disruptant. By fluorescence analysis, it was proposed that CpcG2 protein connects the rods with a minor allophycocyanin component, to support energy transfer to Photosystem I.

Xiao Xing Geng

Papers

Mutational analysis of genes involved in pilus structure, motility and transformation competency in the unicellular motile cyanobacterium Synechocystis sp. PCC 6803.

Novel Putative Photoreceptor and Regulatory Genes Required for the Positive Phototactic Movement of the Unicellular Motile Cyanobacterium Synechocystis sp. PCC 6803

A Eukaryotic-Type Protein Kinase, SpkA, Is Required for Normal Motility of the Unicellular Cyanobacterium Synechocystis sp. Strain PCC 6803

Distinct roles of CpcG1 and CpcG2 in phycobilisome assembly in the cyanobacterium Synechocystis sp. PCC 6803

FUNCTIONAL ANALYSIS OF LEXA-LIKE GENE, sll 1626 IN Synechocystis sp. PCC 6803 USING DNA MICROARRAY