Establishment of an unfed strain of Paramecium bursaria and analysis of associated bacterial communities controlling its proliferation.

The ciliate Paramecium bursaria harbors several hundred symbiotic algae in its cell and is widely used as an experimental model for studying symbiosis between eukaryotic cells. Currently, various types of bacteria and eukaryotic microorganisms are used as food for culturing P. bursaria; thus, the cultivation conditions are not uniform among researchers. To unify cultivation conditions, we established cloned, unfed strains that can be cultured using only sterile medium without exogenous food. The proliferation of these unfed strains was suppressed in the presence of antibiotics, suggesting that bacteria are required for the proliferation of the unfed strains. Indeed, several kinds of bacteria, such as Burkholderiales, Rhizobiales, Rhodospirillales, and Sphingomonadales, which are able to fix atmospheric nitrogen and/or degrade chemical pollutants, were detected in the unfed strains. The genetic background of the individually cloned, unfed strains were the same, but the proliferation curves of the individual P. bursaria strains were very diverse. Therefore, we selected multiple actively and poorly proliferating individual strains and compared the bacterial composition among the individual strains using 16S rDNA sequencing. The results showed that the bacterial composition among actively proliferating P. bursaria strains was highly homologous but different to poorly proliferating strains. Using unfed strains, the cultivation conditions applied in different laboratories can be unified, and symbiosis research on P. bursaria will make great progress.

GPR62 constitutively activates cAMP signaling but is dispensable for male fertility in mice.

G-protein-coupled receptors (GPCRs) participate in diverse physiological functions and are promising targets for drug discovery. However, there are still over 140 orphan GPCRs whose functions remain to be elucidated. Gpr62 is one of the orphan GPCRs that is expressed in the rat and human brain. In this study, we found that Gpr62 is also expressed in male germ cells in mice, and its expression increases along with sperm differentiation. GPR62 lacks the BBXXB and DRY motifs, which are conserved across many GPCRs, and it was able to induce cAMP signaling in the absence of a ligand. These structural and functional features are conserved among mammals, and the mutant analysis of GPR62 has revealed that lacking of these motifs is involved in the constitutive activity. We also found that GPR62 can homooligomerize, but it is not sufficient for its constitutive activity. We further investigated its physiological function by using Gpr62 knockout (Gpr62-/-) mice. Gpr62-/- mice were born normally and did not show any abnormality in growth and behavior. In addition, both male and female Gp62-/- mice were fertile, and the differentiation and motility of spermatozoa were normal. We also found that Gpr61, the gene most similar to Gpr62 in the GPCR family shows a constitutive activity and an expression pattern similar to those of Gpr62 Our results suggest that GPR62 constitutively activates the cAMP pathway in male germ cells but is dispensable for male fertility, which is probably due to its functional redundancy with GPR61.