We have identified by differential screening a novel Arabidopsis thaliana gene, called kin1, which is induced at +4 degrees C. The nucleotide sequences of both the genomic clone and the corresponding cDNA were determined. The deduced 6.5 kDa polypeptide has an unusual amino acid composition being rich in alanine, glycine and lysine. The gene belongs to a family of at least two genes. Northern blot analysis revealed that the level of kin1 mRNA is increased 20-fold in cold-treated plants. In addition to being expressed in cold, kin1 is also induced by water stress and the plant hormone abscisic acid (ABA) which has been suggested to be a common mediator for osmotic stress responses and cold acclimation in plants. Sequence comparisons showed that the kin1 gene product has similarities to fish antifreeze proteins (AFPs).
The lacZ gene of Escherichia coli, coding for beta‐galactosidase, is a widely used reporter gene for gene expression studies in microbial and animal systems. To demonstrate that it is also a powerful reporter gene in plants, lacZ was fused to 5′ regulatory elements of several genes known to be functional in plant cells. By measuring LacZ activities in transgenic plants containing these gene constructs, we showed that the reporter is correctly monitoring the regulatory properties of the well‐characterized promoters fused to lacZ. beta‐Galactosidase was assayed directly in plant extracts when they contained high levels of LacZ or, when LacZ was expressed at low level, by separating the endogenous and LacZ activities electrophoretically and detecting the enzymes with a fluorogenic substrate. The most outstanding property of the marker is its amenability to histochemical detection. Due to its stability, LacZ can be fixed in the tissue with glutaraldehyde without loss of activity and detected with high resolution by using XGal. We could reveal expression patterns unnoticed earlier for many of the regulatory elements studied. The chlorophyll a/b binding protein gene, expressed at very high levels in green tissues, is also expressed at a low level in the vascular cylinder of the root. The Agrobacterium T‐DNA gene encoding octopine synthase is especially active in the epidermis of the root tip and the TR2′ gene was shown to be root specific in the intact plant and stimulated by wounding in the leaf tissue. The TR1′ gene, fused to nptII, shows similar characteristics suggesting co‐regulation of this tightly linked dual promoter.
Exposure of Arabidopsis thaliana L. to an acclimation temperature (+4°C) results in a rapid increase of frost tolerance from -3°C to -7°C. This increase could be correlated to changes in soluble protein pattern. Analysis of in vitro translation products from isolated mRNA suggests that induction acts at the transcriptional level.
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