The differential expression of six basic helix-loop-helix (bHLH) genes in response to low temperatures was studied by evaluating their mRNA levels in the buds and shoots of grapevines. Comparison of the amino acid sequences deduced from nucleotide sequences of the bHLH genes in Vitis labruscana transcripts revealed homologies ranging from 58.8% (VlCEbHLH1 and VlCEbHLH4) to 8.2% (VvbHLH and VlCEbHLH3). Specific primers from unique regions of genes were obtained by alignment of nucleotide sequences and used to evaluate differential expression patterns of bHLH genes. The accumulation patterns of bHLH mRNAs were induced differentially and dependent on the treated temperatures, −20°C in the buds and −2°C and 4°C in the shoots of ‘Campbell Early’ and ‘Muscat Baily A’ grapevines. The bHLH genes showed differential expression patterns in response to low temperatures. In ‘Campbell Early’, the expression of VlCEbHLH1, VlCEbHLH2, and VlCEbHLH5 was up-regulated, while VlCEbHLH3 was down-regulated in response to −20°C cold stress. In ‘Muscat Bailey A’, the expression of all genes was rapidly up-regulated, peaking at 6 h, then decreasing at 12 h after treatment.

Grape (Vitis spp.), which is an important economic fruit crop worldwide, is the fourth most common fruit produced in Korea. Low temperature is one of the most important environmental factors that affect the successful cultivation of grapes. To screen for specific expression of abiotic stress-related genes against cold treatment in ‘Campbell Early’ and ‘Muscat Bailey A’ grapevines, the expression of various defense-related genes was investigated by real-time PCR and semi-quantitative RT-PCR. Among 67 genes analyzed by real-time PCR and RT-PCR, 43 and 10 were found to be up-regulated, while 4 and 37 were down-regulated in low temperature treated ‘Campbell Early’ and ‘Muscat Baily A’ grapevines, respectively. Genes encoding cold induced protein (CIP), lipoxygenase (LOX), glutathione S-transferase (GST), polygalacturonase-inhibiting protein (PGIP), and cold acclimation specific protein (CAS15A) were up-regulated, while those encoding cinnamyl alcohol dehydrogenase 2 (CAD2) and cold acclimation protein (WCS120) were down-regulated by low temperature treatment in both ‘Campbell Early’ and ‘Muscat Bailey A’. These changes in gene expression showed that defense response-related genes for cold acclimation were induced and wheat cold-specific gene decreased in grapevines. Among tested genes, the transcript levels of CLP, cell wall protein (CW), TIP, TLP, and WRKY transcription factor 10 (WRKY10) increased considerably in response to low temperature in ‘Campbell Early’ grapevine. These findings suggest that differentially expressed genes could be utilized as candidates for useful genetic resources and molecular markers after additional confirmation in various grapevine cultivars treated with low temperature in breeding programs for grapes with cold-tolerance.

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