Comparative bioinformatics analysis and abiotic stress responses of expansin proteins in Cucurbitaceae members: watermelon and melon.

Watermelon and melon are members of the Cucurbitaceae family including economically significant crops in the world. The expansin protein family, which is one of the members of the cell wall, breaks down the non-covalent bonds between cell wall polysaccharides, causing pressure-dependent cell expansion. Comparative bioinformatics and molecular characterization analysis of the expansin protein family were carried out in the watermelon (Citrullus lanatus) and melon (Cucumis melo) plants in the study. Gene expression levels of expansin family members were analyzed in leaf and root tissues of watermelon and melon under ABA, drought, heat, cold, and salt stress conditions by quantitative real-time PCR analysis. After comprehensive searches, 40 expansin proteins (22 ClaEXPA, 14 ClaEXPLA, and 4 ClaEXPB) in watermelon and 43 expansin proteins (19 CmEXPA, 15 CmEXPLA, 3 CmEXPB, and 6 CmEXPLB) in melon were identified. The greatest orthologous genes were identified with soybean expansin genes for watermelon and melon. However, the latest divergence time between orthologous genes was determined with poplar expansin genes for watermelon and melon expansin genes. ClaEXPA-04, ClaEXPA-09, ClaEXPB-01, ClaEXPB-03, and ClaEXPLA-13 genes in watermelon and CmEXPA-12, CmEXPA-10, and CmEXPLA-01 genes in melon can be involved in tissue development and abiotic stress response of the plant. The current study combining bioinformatics and experimental analysis can provide a detailed characterization of the expansin superfamily which has roles in growth and reaction to the stress of the plant. The study ensures detailed data for future studies examining gene functions including the roles in plant growth and stress conditions.

Comparative genomic analysis of expansin superfamily gene members in zucchini and cucumber and their expression profiles under different abiotic stresses.

Zucchini and cucumber belong to the Cucurbitaceae family, a group of economical and nutritious food plants that is consumed worldwide. Expansin superfamily proteins are generally localized in the cell wall of plants and are known to possess an effect on cell wall modification by causing the expansion of this region. Although the whole genome sequences of cucumber and zucchini plants have been resolved, the determination and characterization of expansin superfamily members in these plants using whole genomic data have not been implemented yet. In the current study, a genome-wide analysis of zucchini (Cucurbita pepo) and cucumber (Cucumis sativus) genomes was performed to determine the expansin superfamily genes. In total, 49 and 41 expansin genes were identified in zucchini and cucumber genomes, respectively. All expansin superfamily members were subjected to further bioinformatics analysis including gene and protein structure, ontology of the proteins, phylogenetic relations and conserved motifs, orthologous relations with other plants, targeting miRNAs of those genes and in silico gene expression profiles. In addition, various abiotic stress responses of zucchini and cucumber expansin genes were examined to determine their roles in stress tolerance. CsEXPB-04 and CsEXPA-11 from cucumber and CpEXPA-20 and CpEXPLA-14 from zucchini can be candidate genes for abiotic stress response and tolerance in addition to their roles in the normal developmental processes, which are supported by the gene expression analysis. This work can provide new perspectives for the roles of expansin superfamily genes and offers comprehensive knowledge for future studies investigating the modes of action of expansin proteins. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-021-01108-w.

Retinoic acid-stimulated ERK1/2 pathway regulates meiotic initiation in cultured fetal germ cells.

In murine fetal germ cells, retinoic acid (RA) is an extrinsic cue for meiotic initiation that stimulates transcriptional activation of the Stimulated by retinoic acid gene 8 (Stra8), which is required for entry of germ cells into meiotic prophase I. Canonically, the biological activities of RA are mediated by nuclear RA receptors. Recent studies in somatic cells found that RA noncanonically stimulates intracellular signal transduction pathways to regulate multiple cellular processes. In this study, using a germ cell culture system, we investigated (1) whether RA treatment activates any mitogen-activated protein kinase (MAPK) pathways in fetal germ cells at the time of sex differentiation, and (2) if this is the case, whether the corresponding RA-stimulated signaling pathway regulates Stra8 expression in fetal germ cells and their entry into meiosis. When XX germ cells at embryonic day (E) 12.5 were cultured with RA, the extracellular-signal-regulated kinase (ERK) 1/2 pathway was predominantly activated. MEK1/2 inhibitor (U0126) treatment suppressed the mRNA expressions of RA-induced Stra8 and meiotic marker genes (Rec8, Spo11, Dmc1, and Sycp3) in both XX and XY fetal germ cells. Furthermore, U0126 treatment dramatically reduced STRA8 protein levels and numbers of meiotic cells among cultured XX and XY fetal germ cells even in the presence of RA. Taken together, our results suggest the novel concept that the RA functions by stimulating the ERK1/2 pathway and that this activity is critical for Stra8 expression and meiotic progression in fetal germ cells.

Immune responses to methanolic extract of black cumin (Nigella sativa) in rainbow trout (Oncorhynchus mykiss).

The immune stimulating effects of the methanolic extract of black cumin (Nigella sativa) in rainbow trout (Oncorhynchus mykiss) was evaluated. Variable concentrations of black cumin methanolic extract [0 (Control), 0.1 and 0.5 g kg-1 of feed] were individually added to the basal diet and rainbow trout was fed for 30 days to assess the innate immune responses and growth performance. Feed conversion ratio significantly decreased in the group fed with 0.5 g kg-1 black cumin extract. Respiratory burst activity was observed to be the highest in the 0.5 g kg-1 black cumin extract fed group. Lysozyme and myeloperoxidase activities were significantly increased in fish of experimental groups compared to control (P < 0.05). TGF-ß gene expression increased in black cumin 0.5 g kg-1 treated group. IL-1ß and TGF-ß gene expressions decreased in black cumin 0.1 g kg-1 administered group. Expression of IL-12 gene diminished in both the experimental groups. There was no significant difference in survival rates between black cumin extract treated fish groups and control (P > 0.05) after challenged with Aeromonas hydrophila. The results indicate that the methanolic extract of black cumin is a stimulator of some innate humoral immune responses, but it is ineffective for cytokine-related gene trancriptions in rainbow trout.

Dose-dependent functions of fibroblast growth factor 9 regulate the fate of murine XY primordial germ cells.

Male differentiation of primordial germ cells (PGCs) is initiated by the inhibition of entry into meiosis and exposure to male-inducing factor(s), which are regulated by somatic elements of the developing gonad. Fibroblast growth factor 9 (FGF9) produced by pre-Sertoli cells is essential for male gonadal differentiation and also contributes to survival and male differentiation of XY PGCs. However, it is not clear how FGF9 regulates PGC fate. Using a PGC culture system, we identified dose-dependent, fate-determining functions of FGF9 in XY PGCs. Treatment with low levels of FGF9 (0.2 ng/ml) increased expression of male-specific Dnmt3L and Nanos2 in XY PGCs. Conversely, treatment with high levels of FGF9 (25 ng/ml) suppressed male-specific gene expression and stimulated proliferation of XY PGCs. Western blotting showed that low FGF9 treatment enhanced p38 MAPK (mitogen-activated protein kinase) phosphorylation in the same cells. In contrast, high FGF9 treatment significantly stimulated the ERK (extracellular signal-regulated kinase)1/2 signaling pathway in XY PGCs. We investigated the relationship between the ERK1/2 signaling pathway stimulated by high FGF9 and regulation of PGC proliferation. An ERK1/2 inhibitor (U0126) suppressed the PGC proliferation that would otherwise be stimulated by high FGF9 treatment, and increased Nanos2 expression in XY PGCs. Conversely, a p38 MAPK inhibitor (SB202190) significantly suppressed Nanos2 expression that would otherwise be stimulated by low FGF9 in XY PGCs. Taken together, our results suggest that stage-specific expression of FGF9 in XY gonads regulates the balance between proliferation and differentiation of XY PGCs in a dose-dependent manner.

Innate immune and growth promoting responses to caper (Capparis spinosa) extract in rainbow trout (Oncorhynchus mykiss).

Cytokine responses, non-specific immune activity and growth promotion effect of dietary caper (Capparis spinosa) supplementation were examined in rainbow trout (Oncorhynchus mykiss). Rainbow trout (12.04 ± 0.71 g) were fed diets containing three doses of caper methanolic extract [0 (Control), 0.1 and 0.5 g kg(-1) of feed] for 30 days. At the end of the feeding trial, expression levels of cytokine genes that included IL-1ß, IL-8, TGF-ß, IL-12p40, TNF-α1 and IL-10 in head kidney was analyzed using qRT-PCR, and blood and serum were collected to determine superoxide anion production (SAP), phagocytic, lysozyme and myeloperoxidase activities. Expression levels of all cytokines, except TNF-α1 were elevated in the 0.1 g kg(-1) caper extract fed fish group compared to other groups. In 0.5 g kg(-1) caper extract treated fish, only IL-12p40 and IL-10 genes were up-regulated compared to control group fish. SAP was increased in both caper extract treated groups compared to the control, and the highest level was observed in the 0.1 g kg(-1) group. Phagocytic activity in both the caper extract treated groups was increased compared to control with no differences observed between those groups. Lysozyme and myeloperoxidase activities were recorded to be the highest in the 0.1 g kg(-1) fed fish group compared to other groups. Growth promotion was affected positively when caper doses were increased. Survival rate was significantly higher in 0.1 and 0.5 g kg(-1) caper extract treated fish groups compared to control (P < 0.05) after challenged with Aeromonas hydrophila. These results indicate that caper extract stimulates innate immunity through cytokine-mediated responses and promote growth in rainbow trout.