Isolation of the Buchnera aphidicola flagellum basal body complexes from the Buchnera membrane.

Buchnera aphidicola is an intracellular bacterial symbiont of aphids and maintains a small genome of only 600 kbps. Buchnera is thought to maintain only genes relevant to the symbiosis with its aphid host. Curiously, the Buchnera genome contains gene clusters coding for flagellum basal body structural proteins and for flagellum type III export machinery. These structures have been shown to be highly expressed and present in large numbers on Buchnera cells. No recognizable pathogenicity factors or secreted proteins have been identified in the Buchnera genome, and the relevance of this protein complex to the symbiosis is unknown. Here, we show isolation of Buchnera flagellum basal body proteins from the cellular membrane of Buchnera, confirming the enrichment of flagellum basal body proteins relative to other proteins in the Buchnera proteome. This will facilitate studies of the structure and function of the Buchnera flagellum structure, and its role in this model symbiosis.

A genomic reappraisal of symbiotic function in the aphid/Buchnera symbiosis: reduced transporter sets and variable membrane organisations.

Buchnera aphidicola is an obligate symbiotic bacterium that sustains the physiology of aphids by complementing their exclusive phloem sap diet. In this study, we reappraised the transport function of different Buchnera strains, from the aphids Acyrthosiphon pisum, Schizaphis graminum, Baizongia pistaciae and Cinara cedri, using the re-annotation of their transmembrane proteins coupled with an exploration of their metabolic networks. Although metabolic analyses revealed high interdependencies between the host and the bacteria, we demonstrate here that transport in Buchnera is assured by low transporter diversity, when compared to free-living bacteria, being mostly based on a few general transporters, some of which probably have lost their substrate specificity. Moreover, in the four strains studied, an astonishing lack of inner-membrane importers was observed. In Buchnera, the transport function has been shaped by the distinct selective constraints occurring in the Aphididae lineages. Buchnera from A. pisum and S. graminum have a three-membraned system and similar sets of transporters corresponding to most compound classes. Transmission electronic microscopic observations and confocal microscopic analysis of intracellular pH fields revealed that Buchnera does not show any of the typical structures and properties observed in integrated organelles. Buchnera from B. pistaciae seem to possess a unique double membrane system and has, accordingly, lost all of its outer-membrane integral proteins. Lastly, Buchnera from C. cedri revealed an extremely poor repertoire of transporters, with almost no ATP-driven active transport left, despite the clear persistence of the ancestral three-membraned system.

Age- and morph-dependent activation of the lysosomal system and Buchnera degradation in aphid endosymbiosis.

Endosymbiosis in aphids is maintained through a mutualistic association between the host and a symbiotic bacterium, Buchnera, which is harbored in specialized host cells called bacteriocytes. Here, we examined the changes in the Buchnera density in bacteriocytes in relation to the development and polyphenism of the host aphid. Buchnera density in the winged morph aphids, alatae, decreased drastically around the final ecdysis, whereas in the wingless morph aphids, apterae, Buchnera density decreased after the final ecdysis. Thereafter, in both apterae and alatae, Buchnera density was maintained at a constant level until 10 days and then again decreased gradually until 18 days after the final ecdysis. Cytochemical analysis with LysoTracker reagent and quantitative RT-PCR analysis revealed that the number of lysosome-like acidic organelles and the amount of lysosome-related gene (lysozyme and cathepsin L) transcripts increased drastically in the bacteriocytes of alatae around the final ecdysis. Electron microscopy of alatae bacteriocytes around the final ecdysis revealed many Buchnera with irregular electron-dense areas in their cytoplasm that were enclosed by a distended symbiosome membrane. These findings indicated that age- and morph-dependent decreases in Buchnera density coincided with activation of the host lysosomal system and the increased degradation of Buchnera.

Hundreds of flagellar basal bodies cover the cell surface of the endosymbiotic bacterium Buchnera aphidicola sp. strain APS.

Buchnera aphidicola is the endosymbiotic bacterium of the pea aphid. Due to its small genome size, Buchnera lacks many essential genes for autogenous life but obtains nutrients from the host. Although the Buchnera cell is nonmotile, it retains clusters of flagellar genes that lack the late genes necessary for motility, including the flagellin gene. In this study, we show that the flagellar genes are actually transcribed and translated and that the Buchnera cell surface is covered with hundreds of hook-basal-body (HBB) complexes. The abundance of HBB complexes suggests a role other than motility. We discuss the possibility that the HBB complex may serve as a protein transporter not only for the flagellar proteins but also for other proteins to maintain the symbiotic system.