The possible significance of specific biochemical changes in relation to cold acclimation has been discussed extensively (13), but causal relationship between metabolic changes and cold acclimation is difficult to establish. Generally, the development of frost hardiness has been reported to be accompanied with changes in hormones (12,19,24), and in quality and quantity of proteins (14,16). Under a low temperature regime, an increase in leaf soluble protein content was observed only in those potato species which are able to acclimate, i.e. Solanum acaule and Sc4 (4). The net increase in soluble proteins was significantly and positively correlated with the increase of frost hardiness in these species (4). Potato stem cuttings can be easily rooted and propagated in a nutrient agar medium. Such 'stem-cultured plants' can be acclimated to similar frost hardiness levels as with normal tuberpropagated plants grown in pots (5). ABA could substitute for low temperature treatment in inducing frost hardiness in stem-cultured Sc plants, and GA did not affect the cold acclimation (5).The following is a report of experimental results in attempting to determine if there is a relationship between ABA changes and protein synthesis during the development of frost hardiness in the potato during cold acclimation. MATERIALS AND METHODSPlant Materials. Solanum tuberosum L. cv 'Red Pontiac' was propagated from seed tubers, whereas Solanum commersonii (Oka 5040) was propagated from stem cuttings. Plants were planted in 15-cm diameter pots each in a mixture of soil, sand, and peat (3:2:2, v/v). Plants of both species were grown in a uniform environmental chamber for 2 months in a regime of 14-h photoperiod with 450 ,uEm-2 s-' PAR and 20/15°C D/N temperature. Plants of each species were then divided into two groups. One group (controls) was maintained in the same chamber. The second group (treated) was exposed to low temperature of 2°C D/N with a 14-h photoperiod. Frost hardiness of fully expanded leaves was determined after 0, 1, 2, 3, 4, 5, 10, and 15 d of treatments. At the same time, samples were also collected for chemical analyses and moisture determination. The conditions for propagating and growing stem cultures of St and Sc plants were similar to an earlier report (5).Frost Hardiness Evaluation. Excised leaflets or whole shoots (stem-cultured plants) were subjected to controlled freezing (5). After freezing, leaf tissues were placed over ice in a moisturesaturated box in a 5C room and thawed overnight, followed by warming to room temperature. Viability of the tissues after freezing tests was estimated by the conductivity method (20

The effect of temperatures on cold acclimation and deacclimation in foliage tissues was studied in Solan commersonii (Oka 4583), a tuberbearing potato. The threshold temperature for cold acclimation was about 12 C. In a temperature range of 2 to 12 C, the increase in hardiness was dependent on the acclimating temperature; the lower the acclimating temperature, the more hardiness achieved. A day/night temperature of 2 C, regardless of photoperiod, appeared to the optimum acclimating temperature for the SoIuuam species studied. A subfreezing temperature hardened plants less effectively. The maximum level of hardiness could be reached after 15 days of cold acclimation. However, it took only I day to deacclimate the hardened plants to a preacclimation level when plants were subjected to a warm regime from cold. The degree of deacclimation was dependent on the temperature of the warm regime.Based on cold tolerance and the capacity to acclimate to cold, tuberbearing Solawun species could be grouped into five categories. Chilling injury was also observed in some of the tuber-bearing Solanum species.

We have characterized three cDNAs encoding osmotin-like proteins from potato (Solanum commersonii) cell cultures. These cDNAs (pA13, pA35, and pA81) have extensive nucleotide identity in the coding regions but low homology in the 3' non-coding sequences, and may encode three isoforms of potato pathogenesis-related (PR) type 5 proteins. Using gene-specific probes, RNA gel blot analyses showed constitutive accumulation of osmotin-like protein mRNAs in cell cultures, leaves, stems, roots and flowers, with high abundance in the roots and mature flowers. Treatments with abscisic acid (ABA), low temperature, and NaCl increased the accumulation of all three mRNAs in S. commersonii cell cultures and plants grown in vitro. Salicylic acid (SA), and wounding resulted in a moderate increase in the levels of pA13 and pA81 but not pA35 mRNAs. Infection with the fungus Phytophthora infestans activated strong and non-systemic expression of all three osmotin-like protein genes. The accumulation of osmotin-like proteins, however, was detected only in P. infestans-infected tissues but not in plants treated with ABA, SA, NaCl, low temperature, or wounding.

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We have isolated a cDNA (pA13) of an ABA-responsive gene from suspension cultures of Solanum cultures of Solanum commersonii. The deduced amino acid sequence of pA13 cDNA revealed 89 and 91% identity with tobacco osmotin and tomato NP24 protein, respectively. The accumulation of the transcript corresponding to pA13 cDNA was regulated by ABA, cold temperature, and low water potential treatments. Cold-induced accumulation of the pA13 transcript was partially suppressed by fluridone, an ABA synthesis inhibitor, and the suppression was restored by exogenous ABA application. The transcript corresponding to pA13 also accumulated in an organ-specific manner in response to ABA or cold treatment.