Abstract
The ability of organisms to quickly sense and transduce signals of environmental stresses is critical for their survival. Ca2+ is a versatile intracellular messenger involved in sensing a wide variety of stresses and regulating the subsequent cellular responses. So far, our understanding for calcium signaling was mostly obtained from ex vivo tissues and cultured cell lines, and the in vivo spatiotemporal dynamics of stress-triggered calcium signaling in a vertebrate remains to be characterized. Here, we describe the generation and characterization of a transgenic zebrafish line with ubiquitous expression of GCaMP6s, a genetically encoded calcium indicator (GECI). We developed a method to investigate the spatiotemporal patterns of Ca2+ events induced by heat stress. Exposure to heat stress elicited immediate and transient calcium signaling in developing zebrafish. Cells extensively distributed in the integument of the head and body trunk were the first batch of responders and different cell populations demonstrated distinct response patterns upon heat stress. Activity of the heat stress-induced calcium signaling peaked at 30 s and swiftly decreased to near the basal level at 120 s after the beginning of exposure. Inhibition of the heat-induced calcium signaling by LaCl3 and capsazepine and treatment with the inhibitors for CaMKII (Ca²2/calmodulin-dependent protein kinase II) and HSF1 (Heat shock factor 1) all significantly depressed the enhanced heat shock response (HSR). Together, we delineated the spatiotemporal dynamics of heat-induced calcium signaling and confirmed functions of the Ca2+-CaMKII-HSF1 pathway in regulating the HSR in zebrafish.
Highlights
F1 embryos were obtained by mating the positive P0 males with wild type (WT) females and the larvae with green fluorescence were raised to adults
These results indicate that GCaMP6s are ubiquitously expressed, and this transgenic fish model could be used for investigating the in vivo spatiotemporal patterns of calcium signaling triggered by various environmental stressors
These results indicate that GCaMP6s are ubiquitously expressed, 2+ concentrations in the lens and heart suggest that calcium signaling is important high and Ca this transgenic fish model could be used for investigating the in vivo spatiotemporal for visual of perception and hearttriggered functions
Summary
Temperature is the master ecological factor that pervasively affects all cellular processes [1]. Thermal stress can directly disrupt the higher order structures and the functions of macromolecules including proteins, nucleic acids and lipoprotein membranes [2,5]. Metabolic perturbations elicited by thermal stress can generate reactive oxygen species (ROS), which can cause oxidative damages to biomolecules and impair the physiological functions of cells [2,5]. In response to heat stress, cells can initiate a heat shock response (HSR) to minimize the adverse effects and restore cellular homeostasis [3]. HSR is conserved across kingdoms, characterized by rapid and robust induction of the heat shock proteins (HSPs) [2]. HSPs play critical functions in the defense of protein homeostasis, the refolding of denatured proteins, and the breakdown and replacement of the proteins that are unrepairable [3,5,7]
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