Characterization of genetic determinants for R body synthesis and assembly in Caedibacter taeniospiralis 47 and 116

Abstract

Caedibacter taeniospiralis, an obligate bacterial endosymbiont ofParamecium tetraurelia, confers a killing trait upon its host paramecium. Type 51 R bodies (refractile inclusion bodies) are synthesized by these endosymbionts and are required for expression of the killing trait. The nucleotide sequence of the genetic determinants for type 51 R body synthesis and assembly was determined for C. taeniospiralis 47 and 116. Three independently transcribed genes (rebA, rebB, and rebC) were characterized. To date these are the only genes from C. taeniospiralis to be sequenced and characterized. DNA regulatory regions are recognized by Escherichia coli, and codon usage appears similar to that in E. coli. A fourth open reading frame with appropriate regulatory sequences was found within the reb locus, but no evidence was obtained to suggest that this putative gene is expressed in E. coli. The R body-encoding sequences from both strains are identical. Two-dimensional gel electrophoresis of deletion derivatives shows that two polymerization events are involved in R body assembly. One polymerization event requires only RebB and RebC; the other requires all three proteins. Expression of RebC is necessary for the posttranslational modification of RebA and RebB into species with three and two different molecular weights, respectively. In the presence of RebC, each species of RebB with a different molecular weight has six different isoelectric points.Refractile inclusion bodies, known as R bodies, are produced by only a few species of bacteria. These inclusion bodies are highly insoluble protein ribbons, typically seen coiled into cylindrical structures within the cell. Five classes or types of R bodies have been described and are distinguished on the basis of physical dimensions, morphology, and behavior in response to certain physical and chemical treatments (16,21,22).Several classes of R bodies are produced by Caedibacter species, more commonly known as kappa particles. Caedibacter species are obligate bacterial endosymbionts of Paramecium species and are characterized by their ability to produce both R bodies and a toxin. R body-containing forms of Caedibacter species are toxic to sensitive strains of paramecia. Thus, paramecia that carry any Caedibacter strain are referred to as killers. Killing occurs when a sensitive paramecium ingests an R body-containing kappa particle that has been released into the environment by a killer paramecium (6).Five Caedibacter species, producing three of the five known types of R bodies (types 7, 51, and Cc [18,19]), have been identified. R body synthesis in four of the species is determined by extrachromosomal elements thought to be defective phages (17). In the fifth species, Caedibacter taeniospiralis, R body synthesis is directed by plasmid DNA (23). The R bodies of C. taeniospiralis are type 51. They are 0.4 p.m wide, have a maximum length of 20 ,um, possess acute angles at each end, and unroll in a telescopic fashion when exposed to a pH of 6.5 or lower.The R body-encoding region from...

phrR -Like Gene praR of Azorhizobium caulinodans ORS571 Is Essential for Symbiosis with Sesbania rostrata and Is Involved in Expression of reb Genes

“…The paramecia that contain Caedibacter cells producing the R-body kill the paramecia that do not contain endobionts or that contain endobionts that are not the same species of Caedibacter bacteria, and a Caedibacter mutant defective in R-body production does not exhibit killer traits (10,33,34). The rebA, rebB, rebC, and/or rebD genes of C. taeniospiralis are present on plasmids and are involved in R-body synthesis and assembly (20,24,35). Based on these observations, we hypothesized that the phenotype of the A. caulinodans praR mutant was caused by high expression of the reb locus.…”

“…The reb genes were originally identified from C. taeniospiralis (20). Caedibacter species are obligate endobiotic bacteria inhabiting paramecium hosts and are characterized by their ability to produce R-bodies (refractile inclusion bodies), which are insoluble protein ribbons that are seen coiled into cylindrical structures within the cell (33).…”

Lon Protease of Azorhizobium caulinodans ORS571 Is Required for Suppression of reb Gene Expression

“…Furthermore, a Caedibacter mutant that is defective in R-body production has also been observed as not exhibiting "killer traits" (13). The rebA, rebB, rebC, and/or rebD genes of C. taeniospiralis are present on plasmids and are involved in R-body synthesis and assembly (30,35,49). For a long time, the focus of research on reb genes and R-bodies had been only on the relationship between paramecia and Caedibacter bacteria.…”

“…The reb genes were originally identified from Caedibacter taeniospiralis (30). Caedibacter species are obligate endobiotic bacteria inhabiting paramecium hosts and are characterized by their ability to produce R-bodies (refractile inclusion bodies), which are insoluble protein ribbons that are seen coiled into cylindrical structures within the cell (47).…”

The Genus Caedibacter Comprises Endosymbionts of Paramecium spp. Related to the Rickettsiales ( Alphaproteobacteria ) and to Francisella tularensis ( Gammaproteobacteria )

“…This scenario might also explain the otherwise peculiar existence of R-body-like structures in nonsymbiotic bacteria, like Pseudomonas avenae and Pseudomonas taeniospiralis, and their absence in the Caedibacter-related Acanthamoeba endosymbionts (20,23,24,60). While comparative analysis of the genes necessary for R-body production (the genes necessary for type 51 R-body synthesis were cloned and heterologically expressed by Quackenbush and Burbach [19,40]) may help to clarify their evolutionary history, our data clearly demonstrate that the presence of R bodies alone must not be used as a phylogenetically meaningful taxonomic marker.…”