Diethyl ether anesthesia induces transient cytosolic [Ca2+] increase, heat shock proteins, and heat stress tolerance of photosystem II in Arabidopsis

Abstract

General volatile anesthetic diethyl ether blocks sensation and responsive behavior not only in animals but also in plants. Here, using a combination of RNA-seq and proteomic LC–MS/MS analyses, we investigated the effect of anesthetic diethyl ether on gene expression and downstream consequences in plant Arabidopsis thaliana. Differential expression analyses revealed reprogramming of gene expression under anesthesia: 6,168 genes were upregulated, 6,310 genes were downregulated, while 9,914 genes were not affected in comparison with control plants. On the protein level, out of 5,150 proteins identified, 393 were significantly upregulated and 227 were significantly downregulated. Among the highest significantly downregulated processes in etherized plants were chlorophyll/tetrapyrrole biosynthesis and photosynthesis. However, measurements of chlorophyll a fluorescence did not show inhibition of electron transport through photosystem II. The most significantly upregulated process was the response to heat stress (mainly heat shock proteins, HSPs). Using transgenic A. thaliana expressing APOAEQUORIN, we showed transient increase of cytoplasmic calcium level [Ca2+]cyt in response to diethyl ether application. In addition, cell membrane permeability for ions also increased under anesthesia. The plants pre-treated with diethyl ether, and thus with induced HSPs, had increased tolerance of photosystem II to subsequent heat stress through the process known as cross-tolerance or priming. All these data indicate that diethyl ether anesthesia may partially mimic heat stress in plants through the effect on plasma membrane.

Diethyl ether anaesthesia inhibits de‐etiolation of barley seedlings by locking them in intermediate skoto‐photomorphogenetic state

“…This is in line with our previous observations, where we showed the deregulation of many chlorophyll biosynthesis and photosynthesis-related genes in circadian-grown Arabidopsis thaliana after diethyl ether treatment (Pavlovič et al, 2022). This effect can be partially explained by transient diethyl ether-induced increase of cytoplasmic Ca 2+ level [Ca 2+ ] cyt , which is known to be responsible for the repression of LHCB genes through ABI4 (Guo et al, 2016;Pavlovič et al, 2022); ii) second, diethyl ether inhibited de-etiolation, i.e. the light-induced accumulations of chlorophylls and chlorophyll-binding proteins.…”

“…From the data presented here, it is obvious that the effect of diethyl ether is dual: i) first, diethyl ether itself downregulated many genes encoding chlorophyll-binding proteins and enzymes involved in chlorophyll metabolism in the dark as well as in the light. This is in line with our previous observations, where we showed the deregulation of many chlorophyll biosynthesis and photosynthesis-related genes in circadian-grown Arabidopsis thaliana after diethyl ether treatment (Pavlovič et al, 2022). This effect can be partially explained by transient diethyl ether-induced increase of cytoplasmic Ca 2+ level [Ca 2+ ] cyt , which is known to be responsible for the repression of LHCB genes through ABI4 (Guo et al, 2016;Pavlovič et al, 2022); ii) second, diethyl ether inhibited de-etiolation, i.e.…”

“…), (Figure S3, Table S3). Application of diethyl ether (ED/CD and E6/C6 comparisons) induced expression of genes encoding proteins involved in the repair of misfolded proteins (HSP and chaperons) and proteolysis, and reduction of molecular oxygen in accordance with our previous study (Figure S3, Table S3; Pavlovič et al, 2022).…”

“…This down-regulation has been attributed to the release of a plastid factor that indicates a perturbation of the plastid's functional state and represses the expression of nuclear genes required for normal chloro- Verdeja & Strand, 2018;Hernándes-Verdeja et al, 2020). We found inhibited plastid gene expression, but only in dark-grown etiolated seedlings; thus, impaired chlorophyll biosynthesis and ROS production in response to diethyl ether (Table S3; Lin et al, 2011;Yokawa et al, 2018;Pavlovič et al, 2022) are the most prominent candidates. All these suggested mechanisms are not mutually exclusive and all may more or less participate in the inhibited de-etiolation process in barley plants.…”

“…The PIFs transcription factors may act directly or indirectly through another positive regulator, GOLDEN2‐LIKE1 (GLK1) transcription factor (Waters et al, 2009; Martín et al, 2016; Sakuraba et al, 2017). Based on the induction of many stress‐related genes (e.g., HSPs) and ROS production (Table S3; Lin et al, 2011; Yokawa et al, 2018; Pavlovič et al, 2022), the exposition to diethyl ether represents stress conditions for plants. Under such conditions, GLK1 expression is repressed in a GENOME UNCOUPLED1 (GUN1)‐dependent manner (i.e., by retrograde signalling), antagonizing the phytochrome signal and attenuating photomorphogenesis (Martín et al, 2016; Wu and Bock, 2021).…”