Abstract
Many insects are nutritionally dependent on symbiotic microorganisms that have tiny genomes and are housed in specialized host cells called bacteriocytes. The obligate symbiosis between the pea aphid Acyrthosiphon pisum and the γ-proteobacterium Buchnera aphidicola (only 584 predicted proteins) is particularly amenable for molecular analysis because the genomes of both partners have been sequenced. To better define the symbiotic relationship between this aphid and Buchnera, we used large-scale, high accuracy tandem mass spectrometry (nanoLC-LTQ-Orbtrap) to identify aphid and Buchnera proteins in the whole aphid body, purified bacteriocytes, isolated Buchnera cells and the residual bacteriocyte fraction. More than 1900 aphid and 400 Buchnera proteins were identified. All enzymes in amino acid metabolism annotated in the Buchnera genome were detected, reflecting the high (68%) coverage of the proteome and supporting the core function of Buchnera in the aphid symbiosis. Transporters mediating the transport of predicted metabolites were present in the bacteriocyte. Label-free spectral counting combined with hierarchical clustering, allowed to define the quantitative distribution of a subset of these proteins across both symbiotic partners, yielding no evidence for the selective transfer of protein among the partners in either direction. This is the first quantitative proteome analysis of bacteriocyte symbiosis, providing a wealth of information about molecular function of both the host cell and bacterial symbiont.
Highlights
IntroductionWe lack a coherent explanation for the sustained nutrient release from the symbionts and the coordinated growth/proliferation of the two partners, in terms of the metabolic pathways and immune effectors of the host cell and their interactions with the bacteria
Biological Properties of the Fractions Used for Proteome Analysis—We generated a series of samples from pea aphids designed to obtain progressive enrichment of the symbiotic bacterium Buchnera from the whole body of the aphids (WB) through dissected bacteriocytes, partially purified from dissected maternal bacteriocytes (Bu-1) to Buchnera cells purified by Percoll gradient centrifugation (Bu-2) (Fig. 1)
Transmission electron microscopy revealed that the isolated Buchnera cells were intact and were bounded by two membranes (Fig. 1, insert), in contrast to three membranes bounding Buchnera cells in the bacteriocyte [28], suggesting that the outermost membrane, known as the symbiosomal membrane of aphid origin, was eliminated during preparation of Bu-1 and Bu-2
Summary
We lack a coherent explanation for the sustained nutrient release from the symbionts and the coordinated growth/proliferation of the two partners, in terms of the metabolic pathways and immune effectors of the host cell and their interactions with the bacteria. Of particular interest are proteins with known or predicted metabolic or immune function, contributing to nutrient flux among the partners and regulation of the Buchnera cells, respectively. “orphan” proteins (without obvious homologs in other organisms) would suggest specific adaptation of the aphid to its symbiotic relation with Buchnera.
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