Heat Shock Response of Germinating Embryos of Wheat1

Seeds frequently face a hostile environment during early germination. In order to determine whether seeds have evolved unique mechanisms to deal with such environments, a survey of the heat shock response in isolated embryos of wheat (Triticum aestivum L.) was undertaken. Embryos simultaneously heat shocked and labeled following several different periods of prior imbibition up to 12 hours synthesized many groups of heat shock proteins (hsps) typical of other plant and animal systems. Also, five developmentally dependent hsps, present only in treatments imbibed less than 6 hours prior to heat shock, were detected. These proteins have relative molecular masses of 14, 40, 46, 58, and 60 kilodaltons. One of the developmentally dependent hsps is among the most highly labeled hsps found in eardy imbibed embryos. The possibility that this protein is the Em protein is discussed. The hypothesis that the capacity for hsp synthesis is affected by seed vigor was also tested. The heat shock responses of embryos from two high and two low vigor seed lots were compared using oneand two-dimensional electrophoresis of labelled protein extracts. The results indicate that both of the low vigor lots tested had weaker heat shock responses than their high vigor counterparts overall. Not all hsps were relatively less abundant in low vigor embryos. The developmentally dependent hsps showed little relationship to vigor. Some of the developmentally dependent hsps were actually made in greater amounts, relative to other proteins, in the low vigor seed lots. The results presented here demonstrate that imbibing embryos are capable of expressing an enhanced heat shock response, and that this response is related to seed vigor. Upon imbibition, the quiescent seed embryo faces a hostile environment. Extremes of temperature and moisture may confront the young plant simultaneously or in rapid succession. The imbibing embryo must survive this environment in order to germinate. Since the embryo has no developed leaf or root system, it is unable to regulate its temperature and water status through control of transpiration as do mature plants. In addition to a potentially hostile environment, the embryo must deal with the problem of germination itself. Mem' Research was supported in part by National Science Foundation grant RII-8610680 (R. H. A.) and United States Department of Agriculture Hatch Funds to the Wyoming Agricultural Experiment Station. This paper is Wyoming AES number JA1582. branes must go from a disorganized state to the ordered bilayer found in functional cells. Mitochondria, owing in large measure to the disorganization of their membranes, are inactive in the quiescent embryo and must be repaired or replaced in order for germination to occur. Polysomes must be assembled, and new protein and nucleic acid synthesis are initiated immediately upon hydration (6). Osborne (18) has demonstrated that the DNA of quiescent rye embryos accumulates nicks which must also be repaired in order for normal development of the young plant to proceed. Seed vigor is a measure of the seed's ability to germinate and establish under less than optimal conditions. Seed lots which show high germinability in the laboratory may demonstrate poor field emergence. Seed vigor, then, is a manifestation of the ability to survive a series of environmental stresses during germination (1). Deterioration of seed vigor may take place either during maturation on the mother plant or while the seed is in storage. Observed metabolic deficiencies of low vigor seed have been extensively reviewed (1, 19). The net effect ofthe lesions is decreased seed vigor and, ultimately, loss of seed viability. In general, the biochemical sites of seed deterioration can be grouped into the categories of the accumulation ofDNA damage, accumulation of membrane damage, and loss of protein synthetic ability (1, 3, 5). Indeed, protein synthesis may be required to repair membrane damage (5) and has been a subject of interest as a fundamental cause for loss of vigor. One of the more accurate indicators of seed vigor is the accelerated aging test. This test measures a seed's ability to withstand long periods of high temperature and humidity (3). Low vigor seeds show a marked depression in their ability to survive and germinate following exposure to such conditions, while high vigor lots demonstrate a high frequency of germination following the test. A brief, sublethal heat shock induces the production of a number of so called hsps2 in plants (4, 9, 13). Further, such a heat shock has been shown to protect an organism from a second heat dose that would ordinarily be lethal (15). We hypothesized that the hsr during very early germination plays an important role in the survival and eventual germination of seeds under stressful conditions. The conditions of high heat and humidity found in the accelerated aging test, 2Abbreviations: hsp, heat shock protein; hs, heat shock; IEF, isoelectric focusing; ANOVA, analysis of variance; hsr, heat shock response; Em, early metionine protein.