Project description:For RNA-seq experiments, 20 embryos were placed into 5 mL final volume of sterile ERM. Each treatment group were exposed within a 10-minute window. Exposure groups included ERM, 1 µg/L, and 100 µg/L ifosfamide or 10 and 1000 µg/L citalopram (N=5 beakers per treatment). On 7 days post fertilization (dpf), fish were transferred to a labeled and pre-weighed 1.7 mL tube using a p1000 with the tip cut off, then water was completely removed using a p200. The tube was then immediately flash frozen in liquid nitrogen prior to RNA-seq analysis.
Project description:To understand the effects of Hsp60 deficiency in developing vertebrates, we generated CRISPR/Cas9-mediated hspd1 knockout zebrafish lines by targeting exon 2 to induce a frameshift mutation. We selected an allele with a 56 base pair deletion inducing a frameshift mutation leading to loss of protein functions. We examined the proteome changes in zebrafish larvae at 5 days post fertilization (DPF). Wildtype control and hspd1-/- larvae at 5dpf, were analyzed by TMT and nanoLC-MS/MS based proteomcis. For this purpose, we studied five pools from each genotype, and each pool consisted of five larvae.
Project description:To understand the effects of Hsp60 deficiency in developing vertebrates, we generated CRISPR/Cas9-mediated hspd1 knockout zebrafish lines by targeting exon 2 to induce a frameshift mutation. We selected an allele with a 56 base pair deletion inducing a frameshift mutation leading to loss of protein functions. We examined the transcriptome changes in zebrafish larvae at 5 dpf .
Project description:The Liver X Receptors (LXRs) play important roles in multiple metabolic pathways, including fatty acid, cholesterol, carbohydrate and energy metabolism. To expand the knowledge of the functions of LXR signaling during embryonic development, we performed a whole-genome microarray analysis of Lxr target genes in zebrafish larvae treated with either one of the synthetic LXR ligands T0901317 or GW3965. Assessment of the biological processes enriched by differentially expressed genes revealed a prime role for Lxr in regulating lipid metabolic processes, similarly to the function of LXR in mammals. In addition, exposure to the Lxr ligands induced changes in expression of genes in the neural retina and lens of the zebrafish eye, including the photoreceptor guanylate cyclase activators and lens gamma crystallins, suggesting a potential novel role for Lxr in modulating the transcription of genes associated with visual function in zebrafish. The regulation of expression of metabolic genes was phenotypically reflected in an increased absorption of yolk in the zebrafish larvae, and changes in the expression of genes involved in visual perception were associated with morphological alterations in the retina and lens of the developing zebrafish eye. The regulation of expression of both lipid metabolic and eye specific genes was sustained in 1 month old fish. The transcriptional networks demonstrated several conserved effects of LXR activation between zebrafish and mammals, and also identified potential novel functions of Lxr, supporting zebrafish as a promising model for investigating the role of Lxr during development.
Project description:Antidepressants are widely used for treatment of depression. Imipramine, a typical antidepressant, is recently known to have an effect to restore the expression of brain-derived neurotrophic factor gene, which is decreased in depressed patients, by increasing histone acetylation. However, it is largely unknown whether other antidepressants have similar epigenetic effects to change gene expression. To address this question, we examined the effect of five antidepressants (imipramine, citalopram, duloxetine, amitriptyline, mirtazapine) on expression of genes associated with mental and neurodegenetative disorders. We found that 48-hour treatment with imipramine, citalopram, amitriptyline, mirtazapine, but not duloxetine, caused a large number of differentially expressed genes in primary neocortical neurons in mice. Focused on genes relating to neural protection and neuroplasticity, we discovered that amitriptyline not only enhanced the expression of atf3 and cox2 by increasing histone H3 Lys4 trimethylation (H3K4me3), but also protected neurons against oxygen-glucose deprivation and staurosporine-induced apoptosis. Our study will shed a new insight in understanding of the molecular and epigenetic mechanism of antidepressants. Mouse primary neuronal cultures were used to evaluate 48 hour effects of antidepressants. Briefly, cultured neurons at day in vitro 3 (DIV3) were treated with 5µM imipramine, 10 µM citalopram, 0.1 µM duloxetine, 5 µM amitriptylline, 50 µM mirtazepine. These cultures were harvested after 48 hours (DIV5).