Identification and first characterization of 7 Septate junction proteins (SJP) in decapod crabs and their potential involvement in osmoregulation

Abstract

Similar to tight junction proteins found in vertebrates, septate junction proteins (SJPs) serve a significant role in maintaining the epithelial barrier that regulates paracellular transport in invertebrates. Some limited knowledge exists of the cellular localization and function in insects, however, identification, tissue distribution and function of SJPs in crustaceans remains unknown. The current study investigated SJP homologs in three different decapod crabs: Carcinus maenas, Metacarcinus magister, and Geothelphusa dehaani. Seven SJP candidates - Megatrachea-like, Kune-kune-like, Contactin-like, Neurexin-IV-like, Würmchen1-like, Gliotactin-like, and Mesh like- were identified and characterized using transcriptome mining and expression analysis based on sequence homology of known SJPs in insects. One candidate, Würmchen1-like significantly is shorter overall protein length (85 vs. 160 AAs) in compare to insect Würmchen1. A comparative mRNA expression analysis indicated that SJP expression patterns in the branchial epithelia are unique for each crab species, presumably due to a species-specific life style. Further, the study showed that SJPs are highly expressed in epithelial-rich tissues such as the gills and antennal glands of C. maenas, with much lower mRNA expression levels in muscle tissue, providing additional evidence that the investigated proteins are indeed SJPs. Relative mRNA expression of megatrachea-like and kune-like was highest among all SJPs in the anterior gills in C. maenas, regardless whether acclimated to seawater or brackish water. In marine M. magister, würmchen1 like was highest expressed, while kune-like showed highest mRNA expression levels in G. dehaani. When green crabs were acclimated to brackish water, branchial transcript abundance of all but contacin-like and neurexin-IV-like increased, suggesting roles for megatrachea-like, kune kune-like, würmchen1-like, gliotactin-like and mesh-like in protecting for passive ion (e.g. Na+, Cl , Ca2+, and K+) effluxes and/or water influxes. Exposure to high environmental ammonia, HEA (1 mM NH4Cl), resulted in substantial but transient upregulation of all SJPs mRNA except Mesh-like in the posterior gills of hyper-regulating C. maenas. Interestingly, Mesh-like was the only SJP showing upregulation after a HEA exposure of 7 days. Overall, it was demonstrated that the investigated transcripts most likely encode SJPs. However, further studies are necessary to verify their paracellular localization and barrier function.