----------------- INFORMACIÓN GENERAL ------------------- 1. Título del dataset Bioinformatic and gene expression analysis of the somatostatin/cortistatin gene family in the gilthead seabream (Sparus aurata). 2. Autoría: [Rellenar la información de todos los autores siguiendo el siguiente formato. Repetir el esquema, uno para cada autor.] Nombre: Jose Carlos Campos-Sánchez Institución: Universidad de Murcia Correo electrónico: josecarlos.campos@um.es ORCID: 0000-0003-0303-5412 Nombre: Jhon A. Serna‑Duque Institución: Universidad de Murcia Correo electrónico: jhonalberto.sernad@um.es ORCID: 0000-0001-6962-3722 Nombre: Francisco A. Guardiola Institución: Universidad de Murcia Correo electrónico: faguardiola@um.es ORCID: 0000-0002-1018-5446 Nombre: Alberto Cuesta Institución: Universidad de Murcia Correo electrónico: alcuesta@um.es ORCID: 0000-0002-0906-7995 Nombre: María Ángeles Esteban Institución: Universidad de Murcia Correo electrónico: aesteban@um.es ORCID: 0000-0002-6264-1458 3. Fecha de recogida de los datos (fecha única o rango de fechas): [26-01-2024---07-06-2024] 4. Fecha de depósito de los datos: [05-08-2024] 5. Idioma del conjunto de datos: Inglés ------------------------ INFORMACIÓN METODOLÓGICA ------------------------ 1.Descripción de la metodología utilizada para generar el conjunto de datos. 2. Material and methods. 2.1. Bioinformatics analysis. Somatostatin-like gene and protein sequences were searched in the genome-derived protein database (NCBI S. aurata), from which chromosome localization, gene, mRNA, and protein information were extracted from Gene database (Table 1). Exon-intron structure was generated by the Exon-Intron Graphic Maker (http://wormweb.org/exonintron). Genomic synteny was performed in Genomicus 101.01 using the Actinopterygii class as Root species and selecting the PhyloView to observe the conservation of the neighbourhood of the gilthead seabream SSTs genes in the other species according to the gene phylogenetic tree, and the AlignView to align the gene order according to the species phylogenetic tree. PSS sequences were obtained after analysis of the signal peptide cleavage sites with SignalP 6.0 (https://services.healthtech.dtu.dk/). Gilthead seabream somatostatin protein sequences were compared by multiple alignment (Clustal Omega, EMBL-EBI) and visualized by Jalview 2.11. 3D modelling of proteins structure was predicted using Benchling cloud-based platform (https://benchling.com/). Phylogenetic analysis was performed with MEGA software version 11 using the Neighbor-Joining method to infer the evolutionary history. The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (10,000 replicates) were shown next to the branches. The evolutionary distances were computed using the p-distance method, whilst all ambiguous positions were removed for each sequence pair (pairwise deletion option). The name of somatostatin-like genes and proteins were renamed according to the result obtained in phylogenetic and synteny analyses. Finally, mature somatostatin sequences (SST-14) were used to calculate and estimate the physicochemical properties of the peptides with APD3 calculator (Antimicrobial Peptide Database), whilst antimicrobial activity was predicted in CAMPR4 using the algorithm Support Vector Machine (SVM) (Table 2). 2.2. Genetic analysis. 2.2.1. Fish. Specimens (164.17 g ± 14.58 g, 19.50 cm ± 0.33 cm) of the seawater teleost gilthead seabream (S. aurata) were obtained from a local farm (Mazarrón, Spain), randomly redistributed in tanks (450 L) and kept in re-circulating seawater at the Marine Fish Facilities at the University of Murcia (Spain) during a quarantine period of one month. Seawater used for the tanks was prepared by mixing dechlorinated water with concentrated synthetic sea salt (Vibrant Sea) according to the manufacturer instructions. Water temperature was kept at 20 ± 2 °C with a flow rate of 900 L h−1, 28 ‰ salinity, artificial photoperiod of 12 h light to 12 h dark and continuous aeration. Water ammonium and nitrite levels in the tanks were maintained below 0.1 mg L−1 and 0.2 mg L−1, respectively. Fish were fed a commercial diet (Skretting, Spain) at a rate of 2 % body weight day−1 and were kept 24 h without feeding before sampling. The handling of the specimens was always performed in accordance with the Guidelines of the European Union Council (2010/63/UE) and the Ethical Committee of the University of Murcia (Permit Number A13160416). In all cases, fish were sacrificed with 20 mg L-1 of clove oil (Guinama®), completely bled, and rapidly decapitated before sampling. 2.2.2. In vitro exposures. We evaluated the mRNA levels and their regulation in gilthead seabream samples previously obtained and tested [40–43], considering the 3 Rs principle. Constitutive gene expression in naïve conditions was analysed in the whole brain, forebrain, midbrain and hindbrain, skin, gills, heart, liver, intestine, gonad, head-kidney (HK), spleen, thymus, and blood from three healthy and independent fish specimens. Samples were obtained and immediately frozen in TRIzol® Reagent (Life Technologies) and kept at -80º C. For the in vitro experiments, HKLs from five fish were individually isolated (not pooled) and 107 cells mL-1 were incubated in 48- well microtiter plates (Nunc) at 25 ºC, 85 % humidity for 24 h with: culture L-15 medium (control treatment), 10 μg mL-1 phytohemagglutinin (PHA; Sigma-Aldrich), 50 μg mL-1 synthetic unmethylated cytosine-phosphodiester-guanosine oligodeoxynucleotide 1668 (CpG ODN 1668; sequence TCCATGACGTTCCTGATGCT; Eurogentec), 5 μg mL-1 lipopolysaccharide (LPS; Sigma-Aldrich), 5 μg mL-1 concanavalin A (ConA; Sigma-Aldrich), 25 μg mL-1 Polyinosinic:polycytidylic acid (poly I:C; Sigma), 108 UFC mL-1 live Photobacterium damselae subsp. piscicida, 108 UFC mL-1 or Vibrio anguillarum R-82, or 106 tissue culture infective dose (TCID) 50 mL-1 nodavirus (NNV) [40]. Based on the results obtained in this assay, we used samples obtained from another similar experiment in which HKLs individually isolated from other five fish, adjusted to 107 cells mL-1 and incubated in 24- well microtiter plates (Nunc) at 25 ºC, 85 % humidity and 5% CO2 for 24 h with PBS/DMSO diluted in sRPMI medium (control), 1,000 μg mL-1 λ-carrageenan (Sigma) or 2.5 μg mL-1 cantharidin (Sigma) [41]. Regarding the CMC assay, HKLs (n = 3) were incubated for 4 h with the seabream SAF-1 cell line (ECACC 00122301), mock- or NNV-infected during 24 h, as target cells [42,43]. After exposure, HKLs were obtained by centrifugation (400 x g, 10 min, 22 ºC) and stored at -80 ºC until use. 2.2.3. Gene expression by real-time PCR. Total RNA was extracted with PureLink® RNA Mini Kit (Life Technologies) and cDNA synthesized by the SuperScript IV reverse transcriptase (Life Technologies) as indicated by the manufacturer. Real-time PCR was performed using a real-time qPCR with QuantStudio™ Real-Time PCR System Fast (Life Technologies) using SYBR Green PCR Core Reagents (Applied Biosystems). Reaction mixtures were incubated for 10 min at 95 ◦C, followed by 40 cycles of 15 s at 95 ◦C, 1 min at 60 ◦C, and finally 15 s at 95 ◦C, 1 min 60 ◦C and 15 s at 95 ◦C. The gene expression was analysed using the 2−ΔCt method [44], which was performed as described elsewhere [45]. The specificity of the reactions was analysed using samples without cDNA as negative controls. For each mRNA, gene expression was normalized with the geometric mean of ribosomal protein (s18) and elongation factor 1 alfa (ef1a) expression as house-keeping genes. CT values lower than 40 were used for calculations. Samples in which the CT was undetermined the 2−ΔCt value for calculations was assumed as 0. Primers are shown in Table 3. 2.3. Statistical analysis. Figures were processed by GraphPad 8 software. The results were expressed as mean ± standard error of the mean (SEM). Data were analysed by One-way ANOVA (followed by Tukey tests) to determine differences between experimental groups. Normality of the data was previously assessed using a Shapiro-Wilk test and homogeneity of variance was also verified using the Levene test. All statistical analyses were conducted using the computer package SPSS (28.0 version; SPSS Inc., Chicago, IL, USA) for Windows. The level of significance used was p < 0.05 for all statistical tests. 2. Software o instrumentos necesarios para interpretar los datos: [Incluir la versión del software. Si hace falta un software específico de acceso restringido, explicar cómo obtenerlo. Valorar si es posible cambiar el conjunto de datos a un formato abierto (recomendado).] Data analysis: Computer package SPSS (28.0 version; SPSS Inc., Chicago, IL, USA) for Windows. 3. Procedimientos seguidos para asegurar la calidad de los datos Positive and negative controls in each assay. Samples and controls in triplicates to avoid pipetting errors. ------------------------ ESCTRUCTURA DE LOS ARCHIVOS --------------------------- 1. Nombres de archivos [Mencionar todos los archivos incluidos en el conjunto de datos, con el nombre y la extensión (.csv, .pdf, etc.) de cada fichero]. SST-CST - Raw data.xls 2. Formato de los archivos: Libro de Excel 97-2003 (*.xls) ------------------------ MÁS INFORMACIÓN ------------------------ [Incluir cualquier otra información sobre el conjunto de datos que no haya quedado reflejada en esta plantilla y que se considere relevante.]