Proteomic analysis of symbiosome membranes in Cnidaria–dinoflagellate endosymbiosis

Abstract

Symbiosomes are specific intracellular membrane-bound vacuoles containing microalgae in a mutualistic Cnidaria (host)-dinoflagellate (symbiont) association. The symbiosome membrane is originally derived from host plasma membranes during phagocytosis of the symbiont; however, its molecular components and functions are not clear. In order to investigate the protein components of the symbiosome membranes, homogenous symbiosomes were isolated from the sea anemone Aiptasia pulchella and their purities and membrane intactness examined by Western blot analysis for host contaminants and microscopic analysis using various fluorescent probes, respectively. Pure and intact symbiosomes were then subjected to biotinylation by a cell impermeant agent (Biotin-XX sulfosuccinimidyl ester) to label membrane surface proteins. The biotinylated proteins, both Triton X-100 soluble and insoluble fractions, were subjected to 2-D SDS-PAGE and identified by MS using an LC-nano-ESI-MS/MS. A total of 17 proteins were identified. Based on their different subcellular origins and functional categories, it indicates that symbiosome membranes serve as the interface for interaction between host and symbiont by fulfilling several crucial cellular functions such as those of membrane receptors/cell recognition, cytoskeletal remodeling, ATP synthesis/proton homeostasis, transporters, stress responses/chaperones, and anti-apoptosis. The results of proteomic analysis not only indicate the molecular identity of the symbiosome membrane, but also provide insight into the possible role of symbiosome membranes during the endosymbiotic association.

Cell Biology of Cnidarian-Dinoflagellate Symbiosis

“…Rands et al (297) used enzyme cytochemistry to identify ATPase activity associated with both the animal-and dinoflagellate-derived components of the symbiosome membrane complex of the sea anemone A. viridis, providing evidence for the active control of metabolite transfer. Much more recently, though, the symbiosome membrane complex has been successfully isolated via mechanical dis-ruption of the symbiosis and purification of symbiosomal membranes by continuous sucrose gradient centrifugation; this complex can then visualized with confocal and transmission electron microscopy and its purity assessed with Western blot analysis (184,289,368). Isolation of sufficient symbiosomal membrane material potentially allows for identification of membrane transporters, as well as other components of the membrane, by proteomic analysis or the application of heterologous expression techniques (see, e.g., reference 276).…”

Menthol-induced bleaching rapidly and effectively provides experimental aposymbiotic sea anemones (Aiptasiasp.) for symbiosis investigations

“…can be cleared of symbionts, maintained in an aposymbiotic state for extended periods, and successfully re-infected with a variety of Symbiodinium types (Belda-Baillie et al, 2002;Schoenberg and Trench, 1980;Starzak et al, 2014). The ease of experimental manipulation, combined with the rapidly increasing resolution of genomic, proteomic and metabolic databases for Aiptasia sp., makes this symbiotic anthozoan an ideal candidate for biochemical and genetic experiments that will further our understanding of the cellular processes underlying the cnidarian-dinoflagellate symbiosis (Lehnert et al, 2012;Peng et al, 2010;Sunagawa et al, 2009;Weis et al, 2008).…”

Increased susceptibility of algal symbionts to photo-inhibition resulting from the perturbation of coral gastrodermal membrane trafficking

“…However, most phagosomes containing either heat-killed or photosynthetically-impaired Symbiodinium tested positive for Rab-7. These data indicate that development and regulation of endosymbiosis involves highly coordinated membrane trafficking between the host plasma membrane, the phagosome membrane, and the symbiosome membrane [4].…”

“…However, direct evidence showing that membrane trafficking may modulate symbiosis stability is still lacking. Using proteomic analyses, 30% of the symbiosome membrane-associated proteins have been shown to be derived from the plasma membranes in the gastroderm of the sea anemone Aiptasia pulchella [4].…”

“…Specifically, light irradiation treatment will directly affect the structure and composition of the gastrodermal plasma membrane. This change in membrane composition may directly affect the lipid trafficking that modifies the symbiosome membranes, which would then influence physiological processes within Symbiodinium, such as photosynthesis [4]. To investigate whether there is direct interaction between the host membrane and the resident Symbiodinium populations, this study first isolated symbiotic gastrodermal cells (SGCs, i.e., gastroderm cells with intracellular Symbiodinium) from Euphyllia glabrescens using a noninvasive method.…”