The wheat protein kinase gene,TaPK3/, of the PKABA1 subfamily is differentially regulated in greening wheat seedlings

We have identified a new wheat PKABA1/-like protein kinase gene, TaPK3/, that is expressed in greening wheat seedlings. TaPK3 has high sequence homology (97% similarity with some sequence diversity at the 3' end) to the wheat PKABA1 protein kinase mRNA, which is upregulated by cold-temperature treatment, dehydration and abscisic acid (ABA). Use of a TaPK3 gene-specific probe has revealed that TaPK3 is differentially expressed with respect to PKABA1. TaPK3 mRNA accumulates in greening shoot tissue of wheat, but is not affected by dehydration, cold-temperature treatment or ABA. Based on sequence and expression differences, we conclude that expression of the PKABA1/-like protein kinases is not limited to stress responses.     

Field studies on the regulation of abscisic acid content and germinability during grain development of barley: molecular and chemical analysis of pre-harvest sprouting

To investigate whether the regulation of abscisic acid (ABA) content was related to germinability during grain development, two cDNAs for 9-cis-epoxycarotenoid dioxygenase (HvNCED1 and HvNCED2) and one cDNA for ABA 8'-hydroxylase (HvCYP707A1), which are enzymes thought to catalyse key regulatory steps in ABA biosynthesis and catabolism, respectively, were cloned from barley (Hordeum vulgare L.). Expression and ABA-quantification analysis in embryo revealed that HvNCED2 is responsible for a significant increase in ABA levels during the early to middle stages of grain development, and HvCYP707A1 is responsible for a rapid decrease in ABA level thereafter. The change in the embryonic ABA content of imbibing grains following dormancy release is likely to reflect changes in the expression patterns of HvNCEDs and HvCYP707A1. A major change between dormant and after-ripened grains occurred in HvCYP707A1; the increased expression of HvCYP707A1 in response to imbibition, followed by a rapid ABA decrease and a high germination percentage, was observed in the after-ripened grains, but not in the dormant grains. Under field conditions, HvNCED2 showed the same expression level and pattern during grain development in 2002, 2003, and 2004, indicating that HvNCED2 expression is regulated in a growth-dependent manner in the grains. By contrast, HvNCED1 and HvCYP707A1 showed a different expression pattern in each year, indicating that the expression of these genes is affected by environmental conditions during grain development. The varied expression levels of these genes during grain development and imbibition, which would have effects on the activity of ABA biosynthesis and catabolism, might be reflected, in part, in the germinability in field-grown barley.     

Temperature effects on embryonic abscisic acid levels during development of wheat grain dormancy

Abscisic acid (ABA) levels were determined in both the embryo and remaining grain remnant during development of wheat caryopses under temperature conditions which produced either high or low levels of dormancy in mature grain. Higher levels of grain dormancy were produced in grain from plants grown at 15°C as compared to 25°C. In grain grown at 15°C, embryonic ABA levels steadily increased during development, reaching a maximum at stage IV, just before grain desiccation. At 25°C, ABA levels were very high at the earliest stages of embryonic development, but dropped rapidly during maturation. Only small cultivar differences in ABA levels were observed during development at either temperature. In general, higher levels of dormancy in mature grain correlated with prolonged elevation of ABA levels during grain maturation.     

The stress- and abscisic acid-induced barley gene HVA22: developmental regulation and homologues in diverse organisms

Abscisic acid (ABA) induces the expression of a battery of genes in mediating plant responses to environmental stresses. Here we report one of the early ABA-inducible genes in barley (Hordeum vulgare L.), HVA22, which shares little homology with other ABA-responsive genes such as LEA (late embryogenesis-abundant) and RAB (responsive to ABA) genes. In grains, the expression of HVA22 gene appears to be correlated with the dormancy status. The level of HVA22 mRNA increases during grain development, and declines to an undetectable level within 12 h after imbibition of non-dormant grains. In contrast, the HVA22 mRNA level remains high in dormant grains even after five days of imbibition. Treatment of dormant grains with gibberellin (GA) effectively breaks dormancy with a concomitant decline of the level of HVA22 mRNA. The expression of HVA22 appears to be tissue-specific with the level of its mRNA readily detectable in aleurone layers and embryos, yet undetectable in the starchy endosperm. The expression of HVA22 in vegetative tissues can be induced by ABA and environmental stresses, such as cold and drought. Apparent homologues of this barley gene are found in phylogenetically divergent eukaryotic organisms, including cereals, Arabidopsis, Caenorhabitis elegans, man, mouse and yeast, but not in any prokaryotes. Interestingly, similar to barley HVA22, the yeast homologue is also stress-inducible. These observations suggest that the HVA22 and its homologues encode a highly conserved stress-inducible protein which may play an important role in protecting cells from damage under stress conditions in many eukaryotic organisms.     

Temperature effects on embryonic abscisic acid levels during development of wheat grain dormancy

Abscisic acid (ABA) levels were determined in both the embryo and remaining grain remnant during development of wheat caryopses under temperature conditions which produced either high or low levels of dormancy in mature grain. Higher levels of grain dormancy were produced in grain from plants grown at 15°C as compared to 25°C. In grain grown at 15°C, embryonic ABA levels steadily increased during development, reaching a maximum at stage IV, just before grain desiccation. At 25°C, ABA levels were very high at the earliest stages of embryonic development, but dropped rapidly during maturation. Only small cultivar differences in ABA levels were observed during development at either temperature. In general, higher levels of dormancy in mature grain correlated with prolonged elevation of ABA levels during grain maturation.Contribution from USDA-ARS and the College of Agriculture and Home Economics Research Center, Washington State University, scientific paper no. 8901-13.Mention of a specific product name by the United States Department of Agriculture does not constitute an endorsement and does not imply a recommendation over other suitable products.     

Abscisic acid and gibberellic acid-regulated responses of embryos and aleurone layers isolated from dormant and nondormant barley grains

Dormant and nondormant isogenic barley grains were obtained by maturing grains under short day (SD) or long day (LD) growth conditions, respectively. Hormonal responses of isolated embryos and aleurone layers from these grains were studied. Addition of abscisic acid (ABA) reduced germination rate and percentage of embryos, and induced Rab (ABA-responsive) mRNA in aleurone layers from both types of grain. Embryos and aleurone layers from dormant grains responded stronger to ABA than those from nondormant grains. Gibberellic acid (GA₃) increased the germination rate and percentage of embryos from dormant grains and counteracted the ABA-induced inhibition of embryo germination. GA₃ did not affect the amount of Rab mRNA in aleurone layers, suggesting that expression of the Rab gene has no direct correlation with germination. The stronger response of embryos and aleurone layers from dormant grains to ABA may not be explained by higher endogenous ABA levels, but might be due to differences in hormone signal transduction. Aleurone protoplasts from dormant grains had a higher cytosolic pH than those from nondormant grains. To inhibit the ABA-induced Rab mRNA, a much higher concentration of weak acid was required for aleurone layers from dormant grains than for those from nondormant grains. A possible difference in ABA signal transduction between dormant and nondormant grains is discussed.     

Leucine-rich repeat receptor-like kinase1 is a key membrane-bound regulator of abscisic acid early signaling in Arabidopsis

Abscisic acid (ABA) is important in seed maturation, seed dormancy, stomatal closure, and stress response. Many genes that function in ABA signal transduction pathways have been identified. However, most important signaling molecules involved in the perception of the ABA signal or with ABA receptors have not been identified yet. Receptor-like kinase1 (RPK1), a Leu-rich repeat (LRR) receptor kinase in the plasma membrane, is upregulated by ABA in Arabidopsis thaliana. Here, we show the phenotypes of T-DNA insertion mutants and RPK1-antisense plants. Repression of RPK1 expression in Arabidopsis decreased sensitivity to ABA during germination, growth, and stomatal closure; microarray and RNA gel analysis showed that many ABA-inducible genes are downregulated in these plants. Furthermore, overexpression of the RPK1 LRR domain alone or fused with the Brassinosteroid-insensitive1 kinase domain in plants resulted in phenotypes indicating ABA sensitivity. RPK1 is involved in the main ABA signaling pathway and in early ABA perception in Arabidopsis.     

Regulation of dormancy in barley by blue light and after-ripening: effects on abscisic acid and gibberellin metabolism

White light strongly promotes dormancy in freshly harvested cereal grains, whereas dark and after-ripening have the opposite effect. We have analyzed the interaction of light and after-ripening on abscisic acid (ABA) and gibberellin (GA) metabolism genes and dormancy in barley (Hordeum vulgare 'Betzes'). Analysis of gene expression in imbibed barley grains shows that different ABA metabolism genes are targeted by white light and after-ripening. Of the genes examined, white light promotes the expression of an ABA biosynthetic gene, HvNCED1, in embryos. Consistent with this result, enzyme-linked immunosorbent assays show that dormant grains imbibed under white light have higher embryo ABA content than grains imbibed in the dark. After-ripening has no effect on expression of ABA biosynthesis genes, but promotes expression of an ABA catabolism gene (HvABA8'OH1), a GA biosynthetic gene (HvGA3ox2), and a GA catabolic gene (HvGA2ox3) following imbibition. Blue light mimics the effects of white light on germination, ABA levels, and expression of GA and ABA metabolism genes. Red and far-red light have no effect on germination, ABA levels, or HvNCED1. RNA interference experiments in transgenic barley plants support a role of HvABA8'OH1 in dormancy release. Reduced HvABA8'OH1 expression in transgenic HvABA8'OH1 RNAi grains results in higher levels of ABA and increased dormancy compared to nontransgenic grains.     

Synthesis of abscisic Acid-responsive, heat-stable proteins in embryonic axes of dormant wheat grain

Germination of embryonic axes from dormant grain is inhibited by low concentrations of abscisic acid (ABA) compared with axes from nondormant grain. Incubation of dormant grain axes in 0.05 to 50 micromolar ABA caused the prolonged synthesis of a set of heat-stable proteins. Two of these proteins were identified as dehydrins. In nondormant grain axes, 100- to 1000-fold greater ABA concentrations were required to produce similar results. When embryonic axes of dormant wheat (Triticum aestivum) grain were imbibed without ABA, endogenous ABA levels increased 2.5-fold by 4 hours and then gradually declined. Heat-stable proteins were continuously synthesized for at least 18 hours. No increase in endogenous ABA was observed when nondormant grain axes were imbibed. Endogenous ABA levels in nondormant grain axes remained constant at 4 hours and then declined. The nondormant grain axes initially synthesized the heat-stable proteins, but that synthesis disappeared between 8 and 12 hours. These results showing the prolonged synthesis of ABA-responsive, heat-stable proteins by dormant grain axes, demonstrate that biochemical differences occur when axes from dormant compared with nondormant grains are imbibed.     

Effect of grain colour gene (R) on grain dormancy and sensitivity of the embryo to abscisic acid (ABA) in wheat

The level of grain dormancy and sensitivity to ABA of the embryo, a key factor in grain dormancy, were examined in developing grains of a white-grained wheat line, Novosibirskaya 67 (NS-67), and its red-grained near-isogenic lines (ANK-1A to -1D); a red-grained line, AUS 1490, and its white-grained mutant line (EMS-AUS). ANK lines showed higher levels of grain dormancy than NS-67 at harvest maturity. AUS 1490 grain also showed higher dormancy than EMS-AUS grain. These results suggest that the R gene for grain colour can enhance grain dormancy. However, the dormancy effect conferred by the R gene was not large, suggesting that it plays a minor role in the development of grain dormancy. Water extracts of AUS 1490 and EMS-AUS bran contained germination inhibitors equivalent to 1-10 microM ABA, although there was no difference in the amount of inhibitors between AUS 1490 and EMS-AUS. Thus, the grain colour gene of AUS 1490 did not appear to enhance the level of grain dormancy by accumulating germination inhibitors in its bran. Sensitivity to ABA of embryos was higher in grains collected around harvest-maturity for ANK lines and AUS 1490, compared with NS-67 and EMS-AUS. The R gene might enhance grain dormancy by increasing the sensitivity of embryos to ABA.