The externally derived portion of the hyperosmotic shock-activated cytosolic calcium pulse mediates adaptation to ionic stress in suspension-cultured tobacco cells.
The influx of Ca(2+) into the cytosol has long been suggested to serve as a signaling intermediate in the acquisition of tolerance to hyperosmotic and/or salinity stresses. Here we use aequorin-transformed suspension-cultured tobacco cells to directly assess the role of cytosolic calcium (Ca(2+)(cyt)) signaling in salinity tolerance acquisition. Aequorin luminescence recordings and (45)Ca influx measurements using inhibitors of Ca(2+) influx (Gd(3+) and the Ca(2+)-selective chelator EGTA), and modulators of organellar Ca(2+) release (phospholipase C inhibitors U73122 or neomycin) demonstrate that hyperosmolarity, whether imposed by NaCl or by a non-ionic molecule sorbitol, induces a rapid (returning to baseline levels of Ca(2+) within 10 min) and complex Ca(2+)(cyt) pulse in tobacco cells, deriving both from Gd(3+)-sensitive externally derived Ca(2+) influx and from U73122- and neomycin-sensitive Ca(2+) release from an organelle. To determine whether each of the two components of this brief Ca(2+) signal regulate adaptation to hyperosmotic shock, the Ca(2+) pulse was modified by the addition of Gd(3+), U73122, neomycin, or excess Ca(2+), and then cells were treated with salt or sorbitol. After 10 min the cell culture medias were diluted with additional hyperosmotic media to reduce the toxic affects of the modulators, and the growth of cells was measured after 1 week. Gd(3+) treatment reduced growth in salt relative to control cells but not in sorbitol, and exposure to excess Ca(2+) increased growth in salt but not in sorbitol. In contrast, exposure to inhibitors of IP(3) formation had no effect on growth in salt or sorbitol. Therefore, although hyperosmotic treatment stimulates both Ca(2+) influx and Ca(2+) release from an internal Ca(2+) depot, only Ca(2+) influx has a measurable impact on ionic stress tolerance acquisition in tobacco cell suspensions. In contrast, osmoadaptation in these cells appears to occur independent of Ca(2+) signaling.
Cessna
Stephen G
SG
Departments of Biology and Chemistry, Eastern Mennonite University, 1200 Park Road, Harrisonburg, VA 22802, USA. cessnas@emu.edu
Matsumoto
Tracie K
TK
Lamb
Gregory N
GN
Rice
Shawn J
SJ
Hochstedler
Wendy Wenger
WW
D000222:Adaptation, Physiological; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D003600:Cytosol / Q000737:chemistry; D009997:Osmotic Pressure; D012965:Sodium Chloride / Q000494:pharmacology; D013012:Sorbitol / Q000494:pharmacology; D014026:Nicotiana / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D010738:Type C Phospholipases / Q000378:metabolism
D012965:Sodium Chloride; D013012:Sorbitol; D010738:Type C Phospholipases; D002118:Calcium