Genomic Evidence that Methanotrophic Endosymbionts Likely Provide Deep-Sea Bathymodiolus Mussels with a Sterol Intermediate in Cholesterol Biosynthesis

Abstract

Sterols are key cyclic triterpenoid lipid components of eukaryotic cellular membranes, which are synthesized through complex multi-enzyme pathways. Similar to most animals, Bathymodiolus mussels, which inhabit deep-sea chemosynthetic ecosystems and harbor methanotrophic and/or thiotrophic bacterial endosymbionts, possess cholesterol as their main sterol. Based on the stable carbon isotope analyses, it has been suggested that host Bathymodiolus mussels synthesize cholesterol using a sterol intermediate derived from the methanotrophic endosymbionts. To test this hypothesis, we sequenced the genome of the methanotrophic endosymbiont in Bathymodiolus platifrons. The genome sequence data demonstrated that the endosymbiont potentially generates up to 4,4-dimethyl-cholesta-8,14,24-trienol, a sterol intermediate in cholesterol biosynthesis, from methane. In addition, transcripts for a subset of the enzymes of the biosynthetic pathway to cholesterol downstream from a sterol intermediate derived from methanotroph endosymbionts were detected in our transcriptome data for B. platifrons. These findings suggest that this mussel can de novo synthesize cholesterol from methane in cooperation with the symbionts. By in situ hybridization analyses, we showed that genes associated with cholesterol biosynthesis from both host and endosymbionts were expressed exclusively in the gill epithelial bacteriocytes containing endosymbionts. Thus, cholesterol production is probably localized within these specialized cells of the gill. Considering that the host mussel cannot de novo synthesize cholesterol and depends largely on endosymbionts for nutrition, the capacity of endosymbionts to synthesize sterols may be important in establishing symbiont–host relationships in these chemosynthetic mussels.

Dual energy metabolism of the Campylobacterota endosymbiont in the chemosynthetic snail Alviniconcha marisindica

“…RNA probes for FISH were prepared as described previously [47,48]. To synthesize RNA probes, amplifications of 16S rRNA, hydABs, and soxB genes were conducted by PCR using primers, which were designed based on the sequence data from Tables S2 to S3 and using the constructed plasmids as templates; then the amplicons were ligated into a pTA2 vector (Toyobo, Osaka Japan).…”

“…In situ hybridization was carried out as described previously [47,48], with the following modifications. The gill tissues excised from the in situ fixed individuals were refixed in Davidson's solution (10% glycerol, 40% formaldehyde, 30% ethanol, and 30% filtered artificial seawater) at −30°C for 24 h and at 4°C for 16 h, followed by stepwise dehydration in an ethanol series.…”

microRNAs Facilitate Comprehensive Responses of Bathymodiolinae Mussel Against Symbiotic and Nonsymbiotic Bacteria Stimulation

“…Symbiotic MOBs were puri ed using a method reported previously with some modi cations [33]. In short, gill tissue was homogenized on ice and ltered using sterilized gauze and nylon lters, with successive pore sizes of 10, 5, and 3 µm.…”

“…However, the mechanisms controlling the symbiosis between G. platifrons and symbiotic MOBs still remain largely unknown due to the unavailability of cultivable symbionts and accessible mussels. Several methods have been used to harvest symbionts from Bathymodiolinae mussels to date, including enrichment by differential centrifugation and density gradient centrifugation [33][34][35][36]. It has been found that far fewer MOBs can be yielded from density gradient centrifugation compared to differential centrifugation, although their purity is better.…”

Endosymbionts of Metazoans Dwelling in the PACManus Hydrothermal Vent: Diversity and Potential Adaptive Features Revealed by Genome Analysis

“…Nutrient interdependence strengthens the interactions between host and symbionts, leading to strong and stable symbiosis (105,106). Transporters for carbohydrates, lipids, and amino acids were all available in symbionts' genomes, and they even expanded in some symbionts.…”