The complete genome of Teredinibacter turnerae T7901: an intracellular endosymbiont of marine wood-boring bivalves (shipworms).

Here we report the complete genome sequence of Teredinibacter turnerae T7901. T. turnerae is a marine gamma proteobacterium that occurs as an intracellular endosymbiont in the gills of wood-boring marine bivalves of the family Teredinidae (shipworms). This species is the sole cultivated member of an endosymbiotic consortium thought to provide the host with enzymes, including cellulases and nitrogenase, critical for digestion of wood and supplementation of the host's nitrogen-deficient diet. T. turnerae is closely related to the free-living marine polysaccharide degrading bacterium Saccharophagus degradans str. 2-40 and to as yet uncultivated endosymbionts with which it coexists in shipworm cells. Like S. degradans, the T. turnerae genome encodes a large number of enzymes predicted to be involved in complex polysaccharide degradation (>100). However, unlike S. degradans, which degrades a broad spectrum (>10 classes) of complex plant, fungal and algal polysaccharides, T. turnerae primarily encodes enzymes associated with deconstruction of terrestrial woody plant material. Also unlike S. degradans and many other eubacteria, T. turnerae dedicates a large proportion of its genome to genes predicted to function in secondary metabolism. Despite its intracellular niche, the T. turnerae genome lacks many features associated with obligate intracellular existence (e.g. reduced genome size, reduced %G+C, loss of genes of core metabolism) and displays evidence of adaptations common to free-living bacteria (e.g. defense against bacteriophage infection). These results suggest that T. turnerae is likely a facultative intracellular ensosymbiont whose niche presently includes, or recently included, free-living existence. As such, the T. turnerae genome provides insights into the range of genomic adaptations associated with intracellular endosymbiosis as well as enzymatic mechanisms relevant to the recycling of plant materials in marine environments and the production of cellulose-derived biofuels.

Characterization of the mobilization determinants of pAN12, a small replicon from Rhodococcus erythropolis AN12.

Bacteria belonging to the Gram-positive actinomycete species, Rhodococcus erythropolis, are diverse not only in terms of metabolic potentials but the plasmids they encode. It was shown previously that the R. erythropolis AN12 genome harbors a 6.3kb cryptic plasmid called pAN12, which is a member of the pIJ101 family of plasmids. Here we show that pAN12 is conjugatively mobilizable into other rhodococcal strains. A series of plasmid deletion constructs were tested for loss of mobility to identify the pAN12 cis-acting conjugation requirement. In this way, an approximately 700bp region was found to be required for plasmid transmission. A small 61bp element within this region confers mobility to an otherwise non-mobilizable plasmid. Unlike pIJ101, which encodes all necessary factors for transfer, pAN12 mobility is dependent on the presence of an AN12 megaplasmid, pREA400.

Ptpmeg is required for the proper establishment and maintenance of axon projections in the central brain of Drosophila.

Ptpmeg is a cytoplasmic tyrosine phosphatase containing FERM and PDZ domains. Drosophila Ptpmeg and its vertebrate homologs PTPN3 and PTPN4 are expressed in the nervous system, but their developmental functions have been unknown. We found that ptpmeg is involved in neuronal circuit formation in the Drosophila central brain, regulating both the establishment and the stabilization of axonal projection patterns. In ptpmeg mutants, mushroom body (MB) axon branches are elaborated normally, but the projection patterns in many hemispheres become progressively abnormal as the animals reach adulthood. The two branches of MB alpha/beta neurons are affected by ptpmeg in different ways; ptpmeg activity inhibits alpha lobe branch retraction while preventing beta lobe branch overextension. The phosphatase activity of Ptpmeg is essential for both alpha and beta lobe formation, but the FERM domain is required only for preventing alpha lobe retraction, suggesting that Ptpmeg has distinct roles in regulating the formation of alpha and beta lobes. ptpmeg is also important for the formation of the ellipsoid body (EB), where it influences the pathfinding of EB axons. ptpmeg function in neurons is sufficient to support normal wiring of both the EB and MB. However, ptpmeg does not act in either MB or EB neurons, implicating ptpmeg in the regulation of cell-cell signaling events that control the behavior of these axons.

TraA is required for megaplasmid conjugation in Rhodococcus erythropolis AN12.

Pulsed-field gel electrophoresis (PFGE) revealed three previously uncharacterized megaplasmids in the genome of Rhodococcus erythropolis AN12. These megaplasmids, pREA400, pREA250, and pREA100, are approximately 400, 250, and 100kb, respectively, based on their migration in pulsed-field gels. Genetic screening of an AN12 transposon insertion library showed that two megaplasmids, pREA400, and pREA250, are conjugative. Mobilization frequencies of these AN12 megaplasmids to recipient R. erythropolis SQ1 were determined to be approximately 7x10(-4) and 5x10(-4) events per recipient cell, respectively. It is known for other bacterial systems that a relaxase encoded by the traA gene is required to initiate DNA transfer during plasmid conjugation. Sequences adjacent to the transposon insertion in megaplasmid pREA400 revealed a putative traA-like open reading frame. A targeted gene disruption method was developed to generate a traA mutation in AN12, which allowed us to address the role of the traA gene product for Rhodococcus megaplasmid conjugation. We found that the AN12 traA mutant is no longer capable of transferring the pREA400 megaplasmid to SQ1. Furthermore, we confirmed that the conjugation defect was specifically due to the disruption of the traA gene, as pREA400 megaplasmid conjugation defect is restored with a complementing copy of the traA gene.