Identification and characterization of amphibian SLC26A5 using RNA-Seq

Abstract

Background Prestin (SLC26A5) is responsible for acute sensitivity and frequency selectivity in the vertebrate auditory system. Limited knowledge of prestin is from experiments using site-directed mutagenesis or domain-swapping techniques after the amino acid residues were identified by comparing the sequence of prestin to those of its paralogs and orthologs. Frog prestin is the only representative in amphibian lineage and the studies of it were quite rare with only one species identified. Results Here we report a new coding sequence of SLC26A5 for a frog species, Rana catesbeiana (the American bullfrog). In our study, the SLC26A5 gene of Rana has been mapped, sequenced and cloned successively using RNA-Seq. We measured the nonlinear capacitance (NLC) of prestin both in the hair cells of Rana’s inner ear and HEK293T cells transfected with this new coding gene. HEK293T cells expressing Rana prestin showed electrophysiological features similar to that of hair cells from its inner ear. Comparative studies of zebrafish, chick, Rana and an ancient frog species showed that chick and zebrafish prestin lacked NLC. Ancient frog’s prestin was functionally different from Rana. Conclusions We mapped and sequenced the SLC26A5 of the Rana catesbeiana from its inner ear cDNA using RNA-Seq. The Rana SLC26A5 cDNA was 2292 bp long, encoding a polypeptide of 763 amino acid residues, with 40% identity to mammals. This new coding gene could encode a functionally active protein conferring NLC to both frog HCs and the mammalian cell line. While comparing to its orthologs, the amphibian prestin has been evolutionarily changing its function and becomes more advanced than avian and teleost prestin.

Investigation of Gene Networks in Three Components of Immune System Provides Novel Insights into Immune Response Mechanisms against Edwardsiella tarda Infection in Paralichthys olivaceus

“…These analytical techniques enable the determination of gene interactions and the identification of hub genes most closely associated with the subject matter being investigated. In previous research, these two methods have primarily been employed individually, rather than in conjunction with one another, to identify hub genes [38,39]. In the current study, we not only identified key and core genes with high connectivity within each key module but also innovatively ascertained numerous additional hub genes potentially involved in the regulation of P. olivaceus immunity through a combined approach using PPI network and WGCNA analysis.…”

“…To evaluate the accuracy of the RNA-Seq results, we conducted quantitative reverse transcription-polymerase chain reactions (qRT-PCRs). The qRT-PCR protocol followed the methodology described by Wang et al [38].…”