Heat Shock Protein Expression in Leaves of Cabernet Sauvignon

Abstract

Cabernet Sauvignon plants grown at 25°C were heat shocked at 35°C, 40°C, 45°C, or 50°C for four hours, followed by a four-hour incubation at 25°C. A group of non-heat shocked plants was held continuously at 25°C. Proteins from the first fully expanded leaf on the main shoot of each plant were extracted and separated by two-dimensional electrophoresis. The proteins were visualized by silver-staining, or by western immunoblotting with a monoclonal antibody raised against mung bean HSP70. Twelve proteins accumulated following heat shock at 35°C, 41 accumulated following heat shock at 40°C, and 17 accumulated following heat shock at 45°C. Heat shock at 50°C was a lethal treatment and lead to the breakdown of most leaf proteins. Over half of the proteins synthesized following heat shock at 35°C, 40°C, and 45°C were 15-30 kD in size, suggesting that they may be members of the low molecular weight heat shock protein family. Western immunoblots revealed that the antisera recognized two HSP70 isoforms in the control leaves with molecular weights of approximately 70 kD and pIs of 5.86 and 5.90. Neither isoform increased following heat shock at 35°C, but both increased slightly following heat shock at 40°C. A small amount of a third isoform (pI 5.93) was also present following heat shock at 40°C. When the heat shock temperature was 45°C, one HSP70 isoform (pI 5.86) increased dramatically, another (pI 5.90) increased slightly, and the third isoform that was present at 40°C (pI 5.93) was not visible. Expression of HSP70 was largely inhibited when the heat shock temperature was 50°C. Because HSP70 is thought to be induced by the presence of denatured proteins, this suggests that a large number of heat-denatured proteins may accumulate in the cells at 45°C. Heat shock protein synthesis may be related to the increased thermotolerance that has previously been observed following heat shock of Cabernet Sauvignon plants at 40°C for four hours.