Responses to prolonged low-temperature treatment of imbibed seeds (vernalization) were examined in barley (Hordeum vulgare). These occurred in two phases: the perception of prolonged cold, which occurred gradually at low temperatures, and the acceleration of reproductive development, which occurred after vernalization. Expression of the VERNALIZATION1 gene (HvVRN1) increased gradually in germinating seedlings during vernalization, both at the shoot apex and in the developing leaves. This occurred in darkness, independently of VERNALIZATION2 (HvVRN2), consistent with the hypothesis that expression of HvVRN1 is induced by prolonged cold independently of daylength flowering-response pathways. After vernalization, expression of HvVRN1 was maintained in the shoot apex and leaves. This was associated with accelerated inflorescence initiation and with down-regulation of HvVRN2 in the leaves. The largest determinant of HvVRN1 expression levels in vernalized plants was the length of seed vernalization treatment. Daylength did not influence HvVRN1 expression levels in shoot apices and typically did not affect expression in leaves. In the leaves of plants that had experienced a saturating seed vernalization treatment, expression of HvVRN1 was higher in long days, however. HvFT1 was expressed in the leaves of these plants in long days, which might account for the elevated HvVRN1 expression. Long-day up-regulation of HvVRN1 was not required for inflorescence initiation, but might accelerate subsequent stages of inflorescence development. Similar responses to seed vernalization were also observed in wheat (Triticum aestivum). These data support the hypothesis that VRN1 is induced by cold during winter to promote spring flowering in vernalization-responsive cereals.

An understanding of the genetic regulation of low‐temperature (LT) tolerance is a prerequisite for the development of cold tolerant cultivars for high stress regions. Vernalization requirement in winter habit cereals and photoperiod responsiveness in spring habit cereals has been shown to influence expression of the LT tolerance genes. The objective of the present study was to determine the influence of photoperiod response on expression of LT tolerance genes in vernalization requiring winter habit cultivars Norstar and Warrior wheat (Triticum aestivum L. em. Thell) and Kold barley (Hordeum vulgare L.). These cultivars were subjected to 8‐h‐short day (SD) and 20‐h‐long day (LD) photoperiods at cold acclimating temperature (4°C) over a period of 0 to 98 d. Final leaf number (FLN) was determined at intervals throughout the acclimation period to measure vernalization status. Photoperiod sensitivity did not affect vernalization as both SD and LD plants reached vernalization saturation at the same time. However, a significant increase in leaf number and delayed double ridge formation between 49 and 98 d under 4°C SD non‐inductive flowering condition relative to the LD treatments indicated that SD delayed phenological development. Low‐temperature tolerance gene expression as measured by LT50 was influenced before the signal for floral transition as indicated by FLN measurements. Photoperiodic response of SD sensitive winter barley and wheat cultivars was reflected in the level of expression of LT tolerance beginning in the early stages of vernalization and plant development. Subsequent LT acclimation continued for a longer time and to colder temperatures under SD compared to LD. These results support the hypothesis that vernalization and photoperiod responses regulate the expression of LT tolerance genes through their influence on the rate of plant development.

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Vernalization and photoperiod (PP) responses are developmental mechanisms that allow plants to synchronize their growth and reproductive cycles with the seasonal weather changes. Vernalization requirement has been shown to influence the length of time that low-temperature (LT)-induced genes are up-regulated when cereal species are exposed to acclimating temperatures. The objective of the present study was to determine whether expression of LT-induced Wcs and Wcor gene families is also developmentally regulated by PP response. The LT-tolerant, highly short-day (SD)-sensitive barley (Hordeum vulgare L. cv Dicktoo) was subjected to 8-h SD and 20-h long-day PPs at cold-acclimating temperatures over a period of 70 d. A delay in transition from the vegetative to the reproductive stage under SD resulted in an increased level and longer retention of LT tolerance. Similar WCS and WCOR protein homologs were expressed, but levels of expression were much higher in plants acclimated under SD, indicating that the poor LT tolerance of long-day plants was the result of an inability to maintain LT-induced genes in an up-regulated state. These observations indicate that the PP and vernalization genes influence the expression of LT-induced genes in cereals through separate pathways that eventually converge to activate genes controlling plant development. In both instances, the delay in the transition from the vegetative to the reproductive stage produces increased LT tolerance that is sustained for a longer period of time, indicating that the developmental genes determine the duration of expression of LT-induced structural genes.Low-temperature (LT) tolerance in cereals is dependent upon a highly integrated system of structural, regulatory, and developmental genes. In regions with cold winters, vernalization requirement is an important adaptive feature that delays heading by postponing the transition from the vegetative to the reproductive phase. Time sequence studies have shown that the transition from the vegetative to the reproductive growth stage associated with the point of vernalization saturation is also a critical switch that initiates the down-regulation of LT tolerance genes. As a consequence, full expression of LT tolerance only occurs in the vegetative stage, whereas plants in the reproductive phase have a limited ability to cold acclimate (Fowler et al., 1996b; Mahfoozi et al., 2001). Similarly, photoperiod (PP) sensitivity allows plants to maintain LT tolerance for a longer period of time under short-day (SD) compared with long-day (LD) environments (Mahfoozi et al., 2000(Mahfoozi et al., , 2001. In both instances, the delay in the transition from the vegetative to the reproductive stage produces increased LT tolerance that is sustained for a longer period of time.In winter cereals, the expression of several LTinduced genes is positively correlated with the potential of genotypes to develop LT tolerance. Among these are the wcs120, the wcor410, and the wcs19 gene families. The wcs120 family encodes a group of high...

Low-temperature (LT) stress is one of the major limiting factors in cereal production in cold high-altitude mountainous areas of Iran where cereals are exposed to variable periods of temperatures in the vernalization range during the autumn season. Cereals regulate their development through adaptive mechanisms that are responsive to low but nonfreezing temperatures. We exploited a proteomic approach to determine the interrelationship between vernalization fulfillment and expression of low-temperature (LT)-induced protein in most hardy Norstar and semi-hardy Azar2 wheat (Triticum aestivum L. em. Thell). These cultivars were subjected to 12 h of cold acclimating temperature (2 °C) over a period of 0-89 days. LT tolerance, as measured by LT50, and vernalization fulfillment, as estimated from final leaf number (FLN), was determined at intervals throughout the acclimation period. A significant decrease in FLN associated with LT treatment indicated that Norstar and Azar2 had vernalization responses. Azar2 achieved its vernalization fulfillment and maximum LT tolerance (∼ -8 °C) by 28 days of acclimation. However, Norstar had a longer vernalization requirement (between 35 and 42 days) and reached vernalization fulfillment and maximum LT tolerance (∼ -18.7 °C) about the same time as vernalization fulfillment. We applied a two-dimensional electrophoresis-based proteomics approach to analyze changes in the leaf proteome of two genotypes, Norstar and Azar2, during cold acclimation. Using MALDI-TOF/TOF mass spectrometry, 66 LT-associated proteins could significantly be identified. These proteins were categorized into cold-regulated proteins, antifreezing proteins, oxidative stress defense, photosynthesis, chloroplast post-transcriptional regulation, metabolisms, and protein synthesis. A close association between the vernalization fulfillment and the start of a decline in the protein accumulation of hardy Norstar with a long vernalization requirement and semi-hardy Azar2 with a short vernalization requirement was observed. This finding supported the hypothesis that developmental trait which was regulated by vernalization had a regulatory influence over LT proteome response and highlight a close link between the up-regulation of LT-associated proteins and vernalization fulfillment at the molecular level in wheat.