Isolation of four heat shock protein cDNAs from grapefruit peel tissue and characterization of their expression in response to heat and chilling temperature stresses

Abstract

In order to continuously supply horticultural products for long periods, it is essential to store them after harvest in low temperatures. However, many tropical and subtropical fruits and vegetables, such as citrus, are sensitive to chilling. In previous studies, the authors have shown that a short hot water rinsing treatment (at 62degreesC for 20 s) increased chilling tolerance in grapefruit. In order to gain more insight into the molecular mechanisms involved in heat-induced chilling tolerance, PCR cDNA subtraction analysis was performed which isolated four different PCR fragments whose expression was enhanced 24 h after the heat treatment, and that showed high sequence homology with various plant HSP18-I, HSP18-II, HSP22 and HSP70 genes. It was found that the short hot water treatment given at 62degreesC for 20 s, but not at lower temperatures of 20 or 53degreesC, increased the expression of the various HSP cDNAs in grapefruit peel tissue. However, when the fruits were kept at ambient temperatures, the increases in HSP mRNA levels following the hot water treatment were temporary and lasted only between 6 and 48 h. Similar temporary increases in the HSP mRNA levels were detected following exposure of the fruit to a hot air treatment at 40degreesC for 2 h. Nevertheless, when the fruits were treated with hot water but afterwards stored at chilling temperatures of 2degreesC, the mRNA levels of the various HSP18-I, HSP18-II, HSP22 and HSP70 cDNAs increased and remained high and stable during the entire 8-week cold-storage period, suggesting their possible involvement in heat-induced chilling-tolerance responses. The chilling treatment by itself increased the expression of the HSP18-I cDNA, but had no effect on the mRNA levels of any of the other HSP cDNAs. Exposure of fruit to other stresses, such as wounding, UV irradiation, anaerobic conditions and exposure to ethylene, had no effect on the expression of the various HSPs. Overall, the study explored the correlation between the expression and persistence of various HSP cDNAs in grapefruit peel tissue during cold storage, on the one hand, and the acquisition of chilling tolerance, on the other hand, and the results suggest that HSPs may play a general role in protecting plant cells under both high- and low-temperature stresses.