Characterization of Three Related Low-Temperature-Regulated cDNAs from Winter Brassica napus

Abstract

~A cDNA clone, pBNll5, encoding a low-temperature-regulated transcript in winter Brassica napus has been isolated. Northern blot analyses show that levels of transcripts hybridizing to p B N l l 5 increase within 24 h of exposure of B. napus to low temperature, peak at 3 d, and then remain at an elevated level for the duration of the cold treatment (up t o 10 weeks). Transferring plants from 2°C to room temperature results i n the loss of detectable transcripts hybridizing to p B N l l 5 within 1 d. The transcript was not detected in RNA isolated from roots of cold-acclimated B. napus. Results of in vivo labeling of nascent RNA in leaf discs of B. napus with thiouridine suggest that regulation of expression may be transcriptional, at least at the onset of cold temperature. Although drought stress leads t o a slight increase in transcript level at room temperature, neither a brief exposure to elevated temperatures nor exogenous application of abscisic acid resulted in the appearance of the transcript represented by pBNll5. Furthermore, transcripts hybridizing to p B N l l 5 were present at the same levels whether the plants were acclimated in the light or dark. Hybridization experiments show that p B N l l 5 hybridizes strongly to cold-regu- Low temperature is a major trigger for the acquisition of freezing tolerance in plants capable of cold acclimation (Levitt, 1980). Although it has been shown that biochemical, morphological, and physiological changes occur in plant cells during cold acclimation (Sakai and Larcher, 1987; JohnsonFlanagan and Singh, 1988;Singh and Laroche, 1988;Guy, 1990)' direct evidence was obtained only recently to show that low temperature regulated the accumulation of specific mRNAs during cold acclimation (Thomashow, 1990). The appearance of nove1 transcripts during cold acclimation has been observed in alfalfa (Mohapatra et al., 1989), wheat (Lin et al., 1990; Houde et al., 1991), barley (Cattivelli and Bartels, 1990;Dunn et al., 1990), Arabidopsis (Hajela et al., 1990; ' Publication No. 1414 171 Kurkela and Franck, 1990;Nordin et al., 1991; Gilmour et al., 1992), and Brassica (Orr et al., 1992). Furthermore, DNA sequences corresponding to these cold-specific or cold-regulated transcripts have also been isolated and characterized by differential screening of cDNA libraries constructed from these species. These mRNAs appear rapidly upon exposure of the plant to low temperatures, and deduced amino acid sequences of the products of some of these cold-regulated genes have been determined. A number of cold-induced transcripts have been shown to hybridize to transcripts of rab (responsive to ABA) genes (Hahn and Walbot, 1989) or to encode polypeptides containing amino acid sequence motifs (Guo et al., 1991; Gilmour et al., 1992;Houde et al., 1992) found in rab proteins (Skriver and Mundy, 1990), suggesting that they may play a role in the toleration of the cellular desiccation stress that accompanies extracellular freezing. Other cold-induced transcripts encode polypeptides containi...

Constitutive expression of the cold-regulated Arabidopsis thaliana COR15a gene affects both chloroplast and protoplast freezing tolerance

“…Collectively, the independent studies showing that constitutive expression of COR15a affects the freezing tolerance of both chloroplasts frozen in vivo and protoplasts frozen in vitro-together with the extreme reproducibility of the protoplast studies and fact that the enhanced freezing tolerance of chloroplasts and protoplasts occurs over approximately the same temperature range-provide compelling evidence that COR15a is intimately associated with the increase in freezing tolerance that occurs during cold acclimation. Presumably, the coldregulated COR15b gene of A. thaliana (17) and the coldregulated BN115, BN26, and BN19 genes of Brassica napus (33), all of which are apparent homologs of COR15a, also encode polypeptides that affect freezing tolerance.…”

Cold Acclimation and Freezing Tolerance (A Complex Interaction of Light and Temperature)

“…Cold-induced genes have been isolated and characterized in many species, such as alfalfa (Medicago sativa), Arabidopsis tkaliana, barley (Hordeum vulgare), Brassica napus, and wheat (Triticum aestivum) (Houde et al, 1992;Chauvin et al, 1993Chauvin et al, , 1994Dhindsa et al, 1993;Thomashow et al, 1993;Weretilnyk et al, 1993;Danyluk et al, 1994;Crosatti et al, 1995;Limin et al, 1995). There is a high correlation behveen the expression of some of these genes, which appear to be up-regulated by low temperature, and the development of freezing tolerance (Guy, 1990;Thomashow, 1990Thomashow, , 1993Howarth and Ougham, 1993;Lee and Chen, 1993).…”

Expression of a Low-Temperature-Induced Protein in Brassica napus

“…In recent years considerable effort has been directed toward the study of changes in gene expression that occur as a result of cold acclimation. Low-temperature-induced genes have now been described in a number of different plant systems, including wheat (Guo et al, 1992), spinach (Neven et al, 19931, alfalfa (Laberge et al, 1993;Monroy et al, 1993), barley (Cattivelli and Bartels, 1990;Dunn et al, 1994), Arabidopsis (Kurkela and Franck, 1990;Gilmour et al, 1992;Kurkela and Borg-Franck, 1992;Lin and Thomashow, 1992a;Horvath et al, 1993), and Brassica (Orr et al, 1992;Weretilnyk et al, 1993).…”