No abstract

Tobacco (Nicotiana tabacum) and soybean (Glycine max) tissue culture cells were exposed t o a heat shock and protein synthesis studied by SDS-polyacrylamide gel electrophoresis after labeling with radioactive amino acids. A new pattern of protein synthesis is observed in heat-shocked cells compared t o that in control cells. About 12 protein bands, some newly appearing, others synthesized in greatly increased quantities in heat-shock cells, are seen. Several of the heat-shock proteins (HSPs) in both tobacco and soybean are similar in size. One of the HSPs in soybean (76K) shares peptide homology with its presumptive 25°C counterpart, indicating that the synthesis of at least some HSPs may not be due to activation of new genes. The optimum temperature for maximal induction of most HSPs is 39-40°C. Total protein synthesis decreases as heatshock temperature is increased and is barely detectable at 45°C. The heatshock response is maintained for a relatively short time in tobacco cells. After 3 hr at 39"C, a decrease is seen in the synthesis of the HSPs, and after 4 hr practically n o HSPs are synthesized. After exposure t o 39°C for 1 hr, followed by a return of tobacco cells t o 26"C, recovery t o the control pattern of synthesis requires greater than 6 hours. These results indicate that cells of flowering plants exhibit a heat-shock response similar t o that observed in animal cells.Key words: heat-shock, proteins, tobacco, soybeanThe exposure of cells of several different animal species t o heat-shock, ie, a sudden increase in the incubation temperature, results in the inhibition of synthesis of most cell proteins and in the new synthesis of a relatively few proteins. This phenomenon has been extensively described for Drosophila [ 1 , 2 ] , as well as for other insects [3] and in avian and mammalian cells [4].

Soybean seedlings when exposed to a heat shock respond in a manner very similar to that exhibited by cultured cells, and reported earlier [2]. Maximum synthesis of heat shock proteins (HSPs) occurs at 40C. The heat shock response is maintained for a relatively short time under continuous high temperature. After 2.5 hr at 40 C the synthesis of HSPs decreases reaching a very low level by 6 hr. The HSPs synthesized by cultured cells and seedlings are identical and there is a large degree of similarity in HSPs synthesized between the taxonomically widely separated species, soybean and corn. Storage protein synthesis in the developing soybean embryo is not inhibited but is actually stimulated during a heat shock, unlike most other non-HSPs, whose synthesis is greatly reduced. Seedlings respond differently to a gradual increase in temperature than they do a sudden heat shock. There is an upward shift of several degrees in the temperature at which maximum protein synthesis occurs and before it begins to be inhibited. In addition, there appears to be a protection of normal protein synthesis from heat shock inhibition when the temperature increase is gradual. An additional function of the heat shock phenomenon might be the protection of seedlings from death caused by extreme heat stress. The heat shock response appears to have relevance to plants in the field.

The cdc2 kinases are important cell cycle regulators in all eukaryotes. MAP kinases, a closely related family of protein kinases, are involved in cell cycle regulation in yeasts and vertebrates, but previously have not been documented in plants. We used PCR to amplify Brassica napus DNA sequences using primers corresponding to amino sequences that are common to all known protein kinases. One sequence was highly similar to KSS1, a MAP kinase from Saccharomyces cerevisiae. This sequence was used to isolate a full-length MAP kinase-like clone from a pea cDNA library. The pea clone, called D5, shared approximately 50% amino acid identity with MAP kinases from yeasts and vertebrates and about 41% identity with plant cdc2 kinases. An expression protein encoded by D5 was recognized by an antiserum specific to human MAP kinases (ERKs). Messenger RNA corresponding to D5 was present at similar levels in all tissues examined, without regard to whether cell division or elongation were occurring in those tissues.

No abstract