José Muñoz-Dorado

Complete genome sequence of the myxobacterium Sorangium cellulosum.

The genus Sorangium synthesizes approximately half of the secondary metabolites isolated from myxobacteria, including the anti-cancer metabolite epothilone. We report the complete genome sequence of the model Sorangium strain S. cellulosum So ce56, which produces several natural products and has morphological and physiological properties typical of the genus. The circular genome, comprising 13,033,779 base pairs, is the largest bacterial genome sequenced to date. No global synteny with the genome of Myxococcus xanthus is apparent, revealing an unanticipated level of divergence between these myxobacteria. A large percentage of the genome is devoted to regulation, particularly post-translational phosphorylation, which probably supports the strains complex, social lifestyle. This regulatory network includes the highest number of eukaryotic protein kinase–like kinases discovered in any organism. Seventeen secondary metabolite loci are encoded in the genome, as well as many enzymes with potential utility in industry.

Eukaryotic-like protein kinases in the prokaryotes and the myxobacterial kinome

Ser/Thr/Tyr kinases, which together comprise a major class of regulatory proteins in eukaryotes, were not believed to play an important role in prokaryotes until recently. However, our analysis of 626 prokaryotic genomes reveals that eukaryotic-like protein kinases (ELKs) are found in nearly two-thirds of the sequenced strains. We have identified 2697 ELKs, most of which are encoded by multicellular strains of the phyla Proteobacteria (Myxococcales), Actinobacteria, Cyanobacteria, and Chloroflexi, and 2 Acidobacteria and 1 Planctomycetes. Astonishingly, 7 myxobacterial strains together encode 892 ELKs, with 4 of the strains exhibiting a genomic ELK density similar to that observed in eukaryotes. Most myxobacterial ELKs show a modular organization in which the kinase domain is located at the N terminus. The C-terminal portion of the ELKs is highly diverse and often contains sequences with similarity to characterized domains, most of them involved in signaling mechanisms or in protein–protein interactions. However, many of these architectures are unique to the myxobacteria, an observation that suggests that this group exploits sophisticated and novel signal transduction systems. Phylogenetic reconstruction using the kinase domains revealed many orthologous sequence pairs and a huge number of gene duplications that probably occurred after speciation. Furthermore, studies of the microsynteny in the ELK-encoding regions reveal only low levels of synteny among Myxococcus xanthus, Plesiocystis pacifica, and Sorangium cellulosum. However, extensive similarities between M. xanthus, Stigmatella aurantiaca, and 3 Anaeromyxobacter strains were observed, indicating that they share regulatory pathways involving various ELKs.

PomZ, a ParA-like protein, regulates Z-ring formation and cell division in Myxococcus xanthus.

Accurate positioning of the division site is essential to generate appropriately sized daughter cells with the correct chromosome number. In bacteria, division generally depends on assembly of the tubulin homologue FtsZ into the Z‐ring at the division site. Here, we show that lack of the ParA‐like protein PomZ in Myxococcus xanthus resulted in division defects with the formation of chromosome‐free minicells and filamentous cells. Lack of PomZ also caused reduced formation of Z‐rings and incorrect positioning of the few Z‐rings formed. PomZ localization is cell cycle regulated, and PomZ accumulates at the division site at midcell after chromosome segregation but prior to FtsZ as well as in the absence of FtsZ. FtsZ displayed cooperative GTP hydrolysis in vitro but did not form detectable filaments in vitro. PomZ interacted with FtsZ in M. xanthus cell extracts. These data show that PomZ is important for Z‐ring formation and is a spatial regulator of Z‐ring formation and cell division. The cell cycle‐dependent localization of PomZ at midcell provides a mechanism for coupling cell cycle progression and Z‐ring formation. Moreover, the data suggest that PomZ is part of a system that recruits FtsZ to midcell, thereby, restricting Z‐ring formation to this position.

Myxococcus xanthus induces actinorhodin overproduction and aerial mycelium formation by Streptomyces coelicolor

Interaction of the predatory myxobacterium Myxococcus xanthus with the non‐motile, antibiotic producer Streptomyces coelicolor was examined using a variety of experimental approaches. Myxococcus xanthus cells prey on S. coelicolor, forming streams of ordered cells that lyse the S. coelicolor hyphae in the contact area between the two colonies. The interaction increases actinorhodin production by S. coelicolor up to 20‐fold and triggers aerial mycelium production. Other bacteria are also able to induce these processes in S. coelicolor though to a lesser extent. These studies offer new clues about the expression of genes that remain silent or are expressed at low level in axenic cultures and open the possibility of overproducing compounds of biotechnological interest by using potent inducers synthesized by other bacteria.

Copper induction of carotenoid synthesis in the bacterium Myxococcus xanthus

Copper induces a red pigmentation in cells of the bacterium Myxococcus xanthus when they are incubated in the dark, at suboptimal growth conditions. The colouration results from the accumulation of carotenoids, as demonstrated by chemical analysis, and by the lack of a copper effect on M. xanthus mutants affected in known structural genes for carotenoid synthesis. None of several other metals or oxidative agents can mimic the copper effect on carotenoid synthesis. Until now, blue light was the only environmental agent known to induce carotenogenesis in M. xanthus. As happens for the blue light, copper activates the transcription of the structural genes for carotenoid synthesis through the transcriptional activation of the carQRS operon. This encodes the ECF sigma factor CarQ, directly or indirectly responsible for the activation of the structural genes, and the anti‐sigma factor CarR, which physically interacts with CarQ to blocks its action in the absence of external stimuli. All but one of the other regulatory elements known to participate in the induction of carotenoid synthesis by blue light are required for the response to copper. The exception is CarF, a protein required for the light‐mediated dismantling of the CarR–CarQ complex. In addition to carotenogenesis, copper induces other unknown cellular mechanisms that confer tolerance to the metal.

Characterization of manganese-dependent peroxidase isoenzymes from the ligninolytic fungus Phanerochaete flavido-alba.

Phanerochaete flavido-alba is able to decolorize and detoxify olive oil wastewater (OMW) in a process in which simple and polymeric phenols are removed. An unusual acidic MnP is accumulated during the degradation course. This microorganism produces two families of MnPs. MnP1 has an apparent molecular weight of 45 kDa and is secreted as a mixture of isoenzymes with pI ranging from 5.6 to 4.75. MnP2, which is produced as an unique isoenzyme, has an apparent molecular weight of 55.6 Mr and an unusual acidic pI lower than 2.8. The higher specific peroxidase activity for purified MnP2 was for Mn2+ oxidation. Hydroquinone and methylhydroquinone oxidation by MnP2 was Mn2+ dependent, in reaction mixtures without exogenous H2O2. Conversely, ABTS oxidation was Mn2+ independent. Two different DNA fragments (mnpA and mnpB), amplified by PCR, using MnP2 N-terminal sequence and oligonucleotides deduced from two conserved sequences of other MnPs, code for MnPs that belong to the P. chrysosporium mnp2 subfamily on the basis of intron position. The structure of mnpA and mnpB seems to be related to known manganese peroxidase genes, but mnpA encodes an Alanine instead of a Serine (Ser168) regarded as invariant within typical MnPs.

CorE from Myxococcus xanthus Is a Copper-Dependent RNA Polymerase Sigma Factor

The dual toxicity/essentiality of copper forces cells to maintain a tightly regulated homeostasis for this metal in all living organisms, from bacteria to humans. Consequently, many genes have previously been reported to participate in copper detoxification in bacteria. Myxococcus xanthus, a prokaryote, encodes many proteins involved in copper homeostasis that are differentially regulated by this metal. A σ factor of the ECF (extracytoplasmic function) family, CorE, has been found to regulate the expression of the multicopper oxidase cuoB, the P1B-type ATPases copA and copB, and a gene encoding a protein with a heavy-metal-associated domain. Characterization of CorE has revealed that it requires copper to bind DNA in vitro. Genes regulated by CorE exhibit a characteristic expression profile, with a peak at 2 h after copper addition. Expression rapidly decreases thereafter to basal levels, although the metal is still present in the medium, indicating that the activity of CorE is modulated by a process of activation and inactivation. The use of monovalent and divalent metals to mimic Cu(I) and Cu(II), respectively, and of additives that favor the formation of the two redox states of this metal, has revealed that CorE is activated by Cu(II) and inactivated by Cu(I). The activation/inactivation properties of CorE reside in a Cys-rich domain located at the C terminus of the protein. Point mutations at these residues have allowed the identification of several Cys involved in the activation and inactivation of CorE. Based on these data, along with comparative genomic studies, a new group of ECF σ factors is proposed, which not only clearly differs mechanistically from the other σ factors so far characterized, but also from other metal regulators.

Differential Expression of the Three Multicopper Oxidases from Myxococcus xanthus

Myxococcus xanthus is a soil bacterium that undergoes a unique life cycle among the prokaryotes upon starvation, which includes the formation of macroscopic structures, the fruiting bodies, and the differentiation of vegetative rods into coccoid myxospores. This peculiarity offers the opportunity to study the copper response in this bacterium in two different stages. In fact, M. xanthus vegetative rods exhibit 15-fold-greater resistance against copper than developing cells. However, cells pre-adapted to this metal reach the same levels of resistance during both stages. Analysis of the M. xanthus genome reveals that many of the genes involved in copper resistance are redundant, three of which encode proteins of the multicopper oxidase family (MCO). Each MCO gene exhibits a different expression profile in response to external copper addition. Promoters of cuoA and cuoB respond to Cu(II) ions during growth and development; however, they show a 10-fold-increased copper sensitivity during development. The promoter of cuoC shows copper-independent induction upon starvation, but it is copper up-regulated during growth. Phenotypic analyses of deletion mutants reveal that CuoB is involved in the primary copper-adaptive response; CuoA and CuoC are necessary for the maintenance of copper tolerance; and CuoC is required for normal development. These roles seem to be carried out through cuprous oxidase activity.

Myxobacteria: Moving, Killing, Feeding, and Surviving Together

Myxococcus xanthus, like other myxobacteria, is a social bacterium that moves and feeds cooperatively in predatory groups. On surfaces, rod-shaped vegetative cells move in search of the prey in a coordinated manner, forming dynamic multicellular groups referred to as swarms. Within the swarms, cells interact with one another and use two separate locomotion systems. Adventurous motility, which drives the movement of individual cells, is associated with the secretion of slime that forms trails at the leading edge of the swarms. It has been proposed that cellular traffic along these trails contributes to M. xanthus social behavior via stigmergic regulation. However, most of the cells travel in groups by using social motility, which is cell contact-dependent and requires a large number of individuals. Exopolysaccharides and the retraction of type IV pili at alternate poles of the cells are the engines associated with social motility. When the swarms encounter prey, the population of M. xanthus lyses and takes up nutrients from nearby cells. This cooperative and highly density-dependent feeding behavior has the advantage that the pool of hydrolytic enzymes and other secondary metabolites secreted by the entire group is shared by the community to optimize the use of the degradation products. This multicellular behavior is especially observed in the absence of nutrients. In this condition, M. xanthus swarms have the ability to organize the gliding movements of 1000s of rods, synchronizing rippling waves of oscillating cells, to form macroscopic fruiting bodies, with three subpopulations of cells showing division of labor. A small fraction of cells either develop into resistant myxospores or remain as peripheral rods, while the majority of cells die, probably to provide nutrients to allow aggregation and spore differentiation. Sporulation within multicellular fruiting bodies has the benefit of enabling survival in hostile environments, and increases germination and growth rates when cells encounter favorable conditions. Herein, we review how these social bacteria cooperate and review the main cell–cell signaling systems used for communication to maintain multicellularity.

Bacterial predation: 75 years and counting!

The first documented study on bacterial predation was carried out using myxobacteria three quarters of a century ago. Since then, many predatory strains, diverse hunting strategies, environmental consequences and potential applications have been reported by groups all over the world. Now we know that predatory bacteria are distributed in a wide variety of environments and that interactions between predatory and non-predatory populations seem to be the most important factor in bacterial selection and mortality in some ecosystems. Bacterial predation has now been proposed as an evolutionary driving force. The structure and diversity of the predatory bacterial community is beginning to be recognized as an important factor in biodiversity due to its potential role in controlling and modelling bacterial populations in the environment. In this paper, we review the current understanding of bacterial predation, going over the strategies used by the main predatory bacteria to kill their prey. We have also reviewed and integrated the accumulated advances of the last 75 years with the interesting new insights that are provided by the analyses of genomes, predatomes, predatosomes and other comparative genomics studies, focusing on potential applications that derive from all of these areas of study.