Hyperosmotic stress leads to rapid perturbation of ATP levels: a possible role in osmosensing

Debbie McLaggan, Cristina Lagido, Lesley Glover

Research output: Contribution to conferenceAbstract

Abstract

We are interested in the early events in Caenorhabditis elegans following hyperosmotic stress. The molecular mechanisms by which animal cells detect volume changes and transduce those signals into effector responses still remain to be elucidated despite significant progress in the understanding of effector responses. We have established a reproducible, quantitative and scalable bioluminescent assay for the metabolic effects of the first few minutes at high osmolarity. We use our highly luminescent transgenic C. elegans strain PE255 (feIs5) X constructed through the constitutive and generalised expression of the bioluminescent enzyme firefly luciferase (1). We have previously demonstrated that light levels provide a sensitive measure of the ATP levels obtained by RNAi silencing of respiratory chain genes (1). In the context of the study of stress response, luminescence allows effective detection of transient changes in ATP pools in C. elegans that were previously not measureable. Following a hyperosmotic shock, we have observed a transient increase in luminescence of our biosenosr C. elegans, indicative of a transient increase in ATP. The increase in luminescence did not occur in the presence of the respiratory inhibitor sodium azide, indicating that de novo ATP synthesis is required for the observed increase. In addition there was no increased uptake of bioluminescence components upon osmotic upshock as determined by monitoring pharynx pumping. We have determined that the extent of light output is proportional to the extent of osmotic perturbation. This effect also appears to be independent of the initial osmolarity of the medium. Following water loss, the earliest subsequent event that has been determined is the dose dependent increase in WNK1 (with-no-K (lysine) Kinase-1) activation in mammalian cells (2). WNK kinases have important roles in regulating salt and water transport in the mammalian kidney. C. elegans has WNK-1 and other downstream homologues that are required for acute volume recovery in C. elegans (3). These have been postulated to function in a manner similar to mammalian WNK1 kinase signalling pathway (4). The ultimate aim of our studies is to determine whether the transient increase in intracellular ATP is involved in WNK mediated signaling of hyperosmotic water loss. (1) Lagido, C. et. al. (2008) BMC Physiology 8 (1):7. (2) Zagorska, A. et. al. (2007) J. Cell Biol. 1176:89-100. (3) Choe, K.P. and Strange, K. (2007) Am. J. Physiol. 293:C915-C927. (4) Choe, K.P. and Strange, K. (2007) FEBS J. 274:5782-5789.
Original languageEnglish
Publication statusPublished - Jun 2009
Event17th International C. elegans Meeting - Los Angeles, United States
Duration: 24 Jun 200928 Jun 2009

Conference

Conference17th International C. elegans Meeting
CountryUnited States
CityLos Angeles
Period24/06/0928/06/09

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Caenorhabditis elegans
Adenosine Triphosphate
Luminescence
Phosphotransferases
Osmolar Concentration
Water
Luminescent Measurements
Firefly Luciferases
Light
Sodium Azide
Electron Transport
Pharynx
RNA Interference
Cell Size
Lysine
Shock
Salts
Kidney
Enzymes
Genes

Cite this

McLaggan, D., Lagido, C., & Glover, L. (2009). Hyperosmotic stress leads to rapid perturbation of ATP levels: a possible role in osmosensing. Abstract from 17th International C. elegans Meeting, Los Angeles, United States.

Hyperosmotic stress leads to rapid perturbation of ATP levels : a possible role in osmosensing. / McLaggan, Debbie; Lagido, Cristina; Glover, Lesley.

2009. Abstract from 17th International C. elegans Meeting, Los Angeles, United States.

Research output: Contribution to conferenceAbstract

McLaggan, D, Lagido, C & Glover, L 2009, 'Hyperosmotic stress leads to rapid perturbation of ATP levels: a possible role in osmosensing', 17th International C. elegans Meeting, Los Angeles, United States, 24/06/09 - 28/06/09.
McLaggan D, Lagido C, Glover L. Hyperosmotic stress leads to rapid perturbation of ATP levels: a possible role in osmosensing. 2009. Abstract from 17th International C. elegans Meeting, Los Angeles, United States.
McLaggan, Debbie ; Lagido, Cristina ; Glover, Lesley. / Hyperosmotic stress leads to rapid perturbation of ATP levels : a possible role in osmosensing. Abstract from 17th International C. elegans Meeting, Los Angeles, United States.
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N2 - We are interested in the early events in Caenorhabditis elegans following hyperosmotic stress. The molecular mechanisms by which animal cells detect volume changes and transduce those signals into effector responses still remain to be elucidated despite significant progress in the understanding of effector responses. We have established a reproducible, quantitative and scalable bioluminescent assay for the metabolic effects of the first few minutes at high osmolarity. We use our highly luminescent transgenic C. elegans strain PE255 (feIs5) X constructed through the constitutive and generalised expression of the bioluminescent enzyme firefly luciferase (1). We have previously demonstrated that light levels provide a sensitive measure of the ATP levels obtained by RNAi silencing of respiratory chain genes (1). In the context of the study of stress response, luminescence allows effective detection of transient changes in ATP pools in C. elegans that were previously not measureable. Following a hyperosmotic shock, we have observed a transient increase in luminescence of our biosenosr C. elegans, indicative of a transient increase in ATP. The increase in luminescence did not occur in the presence of the respiratory inhibitor sodium azide, indicating that de novo ATP synthesis is required for the observed increase. In addition there was no increased uptake of bioluminescence components upon osmotic upshock as determined by monitoring pharynx pumping. We have determined that the extent of light output is proportional to the extent of osmotic perturbation. This effect also appears to be independent of the initial osmolarity of the medium. Following water loss, the earliest subsequent event that has been determined is the dose dependent increase in WNK1 (with-no-K (lysine) Kinase-1) activation in mammalian cells (2). WNK kinases have important roles in regulating salt and water transport in the mammalian kidney. C. elegans has WNK-1 and other downstream homologues that are required for acute volume recovery in C. elegans (3). These have been postulated to function in a manner similar to mammalian WNK1 kinase signalling pathway (4). The ultimate aim of our studies is to determine whether the transient increase in intracellular ATP is involved in WNK mediated signaling of hyperosmotic water loss. (1) Lagido, C. et. al. (2008) BMC Physiology 8 (1):7. (2) Zagorska, A. et. al. (2007) J. Cell Biol. 1176:89-100. (3) Choe, K.P. and Strange, K. (2007) Am. J. Physiol. 293:C915-C927. (4) Choe, K.P. and Strange, K. (2007) FEBS J. 274:5782-5789.

AB - We are interested in the early events in Caenorhabditis elegans following hyperosmotic stress. The molecular mechanisms by which animal cells detect volume changes and transduce those signals into effector responses still remain to be elucidated despite significant progress in the understanding of effector responses. We have established a reproducible, quantitative and scalable bioluminescent assay for the metabolic effects of the first few minutes at high osmolarity. We use our highly luminescent transgenic C. elegans strain PE255 (feIs5) X constructed through the constitutive and generalised expression of the bioluminescent enzyme firefly luciferase (1). We have previously demonstrated that light levels provide a sensitive measure of the ATP levels obtained by RNAi silencing of respiratory chain genes (1). In the context of the study of stress response, luminescence allows effective detection of transient changes in ATP pools in C. elegans that were previously not measureable. Following a hyperosmotic shock, we have observed a transient increase in luminescence of our biosenosr C. elegans, indicative of a transient increase in ATP. The increase in luminescence did not occur in the presence of the respiratory inhibitor sodium azide, indicating that de novo ATP synthesis is required for the observed increase. In addition there was no increased uptake of bioluminescence components upon osmotic upshock as determined by monitoring pharynx pumping. We have determined that the extent of light output is proportional to the extent of osmotic perturbation. This effect also appears to be independent of the initial osmolarity of the medium. Following water loss, the earliest subsequent event that has been determined is the dose dependent increase in WNK1 (with-no-K (lysine) Kinase-1) activation in mammalian cells (2). WNK kinases have important roles in regulating salt and water transport in the mammalian kidney. C. elegans has WNK-1 and other downstream homologues that are required for acute volume recovery in C. elegans (3). These have been postulated to function in a manner similar to mammalian WNK1 kinase signalling pathway (4). The ultimate aim of our studies is to determine whether the transient increase in intracellular ATP is involved in WNK mediated signaling of hyperosmotic water loss. (1) Lagido, C. et. al. (2008) BMC Physiology 8 (1):7. (2) Zagorska, A. et. al. (2007) J. Cell Biol. 1176:89-100. (3) Choe, K.P. and Strange, K. (2007) Am. J. Physiol. 293:C915-C927. (4) Choe, K.P. and Strange, K. (2007) FEBS J. 274:5782-5789.

M3 - Abstract

ER -