Phytochrome- and Gibberellin-Mediated Regulation of Abscisic Acid Metabolism during Germination of Photoblastic Lettuce Seeds

Abstract

Germination of lettuce (Lactuca sativa) 'Grand Rapids' seeds is regulated by phytochrome. The action of phytochrome includes alterations in the levels of gibberellin (GA) and abscisic acid (ABA). To determine the molecular mechanism of phytochrome regulation of ABA metabolism, we isolated four lettuce cDNAs encoding 9-cis-epoxycarotenoid dioxygenase (biosynthesis; LsNCED1-LsNCED4) and four cDNAs for ABA 8#-hydroxylase (catabolism; LsABA8ox1-LsABA8ox4). Measurements of ABA and its catabolites showed that a decrease in ABA level coincided with a slight increase in the level of the ABA catabolite phaseic acid after red light treatment. Quantitative reverse transcription-polymerase chain reaction analysis indicated that ABA levels are controlled by phytochrome through down-regulation of LsNCED2 and LsNCED4 expression and up-regulation of LsABA8ox4 expression in lettuce seeds. Furthermore, the expression levels of LsNCED4 decreased after GA 1 treatment, whereas the levels of expression of the other two genes were unaffected. The LsNCED4 expression was also down-regulated by red light in lettuce seeds in which GA biosynthesis was suppressed by AMO-1618, a specific GA biosynthesis inhibitor. These results indicate that phytochrome regulation of ABA metabolism is mediated by both GA-dependent and -independent mechanisms. Spatial analysis showed that after red light treatment, the ABA decrease on the hypocotyl side was greater than that on the cotyledon side of lettuce seeds. Moreover, phytochrome-regulated expression of ABA and GA biosynthesis genes was observed on the hypocotyl side, rather than the cotyledon side, suggesting that this regulation occurs near the photoperceptive site.Plants undergo correct morphogenesis by responding to light and adapt to various forms of light. In addition to imbibition, some seeds require light to germinate and are called photoblastic seeds. Photoblastic seeds germinate after irradiation with red light (R; wavelength approximately 600-700 nm), and this effect of R is cancelled by successive irradiation with far-red light (FR; wavelength approximately 700-750 nm). The effects of R and FR are reversible. This type of germination, called photogermination, was discovered by experiments using lettuce (Lactuca sativa) 'Grand Rapids' seeds (Borthwick et al., 1952). The energy from wavelengths of R is mainly required for photosynthesis. This strategy allows survival of small seeds in which storage substances are low, such as the seeds of lettuce, tomato (Solanum lycopersicum), tobacco (Nicotiana tabacum), and Arabidopsis (Arabidopsis thaliana). The typical reversible photoreaction of lettuce seeds allowed Butler et al. (1959) to detect the pigment that acts as the R/FR receptor in extracts from etiolated cotyledons of Brassica rapa and etiolated shoots of Zea mays using differential spectrophotometry, and this photoreceptor was named phytochrome.Many studies to elucidate the mechanisms of photoblastic lettuce seed germination have been carried out for over a half century, including biochemica...

Expression of9-cis-EPOXYCAROTENOID DIOXYGENASE4Is Essential for Thermoinhibition of Lettuce Seed Germination but Not for Seed Development or Stress Tolerance

“…In lettuce, four different NCED genes were identified (Sawada et al, 2008), but their specific expression patterns have not been reported. During seed development, NCED1 was expressed first, followed by NCED2 and then NCED4, while expression of NCED3 was much lower throughout (Figure 4).…”

“…In mature lettuce seeds, NCED4 mRNA is present in the embryo, but not in the endosperm ; information on NCED4 expression during lettuce seed development has not been reported. We therefore assayed mRNA dynamics in developing seeds of the four NCED genes identified in lettuce (NCED1-4) (Sawada et al, 2008). NCED2 expression increased first during seed development and then declined ( Figure 4B), followed by a transient increase in NCED1 expression to two-to threefold higher levels ( Figure 4A).…”

“…In control seeds in which germination was inhibited, mRNA abundance of a number of genes either declined somewhat or was constant following imbibition, but remained elevated after 48 h at 35°C. Among these were Ls-ZEP1 ( Figure 3B), a homolog of the Arabidopsis ABA1 gene encoding an ABA biosynthetic enzyme (Seo and Koshiba, 2002); Ls-ABA8ox4 ( Figure 3C), a homolog of the Arabidopsis CYP707A2 gene encoding an ABA 8'-hydroxlase that initiates ABA catabolism Sawada et al, 2008); transcription factors involved in the ABA signal transduction pathway regulating germination (e.g., Ls-ABI5 and Ls-ABI3; Figures 3D and 3E) (Finkelstein et al, 2008); Ls-SNF4 ( Figure 3F), an ABA-induced gene encoding a regulatory subunit of the SnRK1 kinase family involved in metabolic regulation in seeds (Bradford et al, 2003;Radchuk et al, 2006;Rosnoblet et al, 2007); Ls-GA2ox1 ( Figure 3H), encoding a GA catabolic enzyme (Toh et al, 2008); Ls-LEA ( Figure 3L), encoding a late embryogenesis abundant protein (Delseny et al, 2001); SPATULA ( Figure 3M), a temperature-dependent germination repressor (Penfield et al, 2005); and Ls-DOG1 ( Figure 3O), a homolog of Arabidopsis DOG1, which is involved in the induction and maintenance of seed dormancy (Bentsink et al, 2006; Footitt et al, 2011;Kendall et al, 2011). All of these genes exhibited a tendency for a more rapid decrease in transcript abundance in the RNAi seeds following imbibition, and their mRNAs were very low or absent by 48 h when germination had been completed.…”

Physiological, morphological and biochemical characteristics of the sexual propagation of Piper aduncum (Piperaceae)

“…, Yamaguchi 2008, Seo et al 2009). Perhaps the seeds of P. aduncum are not responsive to the application of GA 3 but to other gibberellins, such as GA 1 as in lettuce seeds (Sawada et al 2008) or GA 4 and GA 7 as in seeds of Taxus mairei (Chien et al 1998). The application of GA 3 negatively influenced the root protrusion of P. aduncum (figure 20).…”

Regulation of abscisic acid metabolism in relation to the dormancy and germination of cereal grains

“…Light quality is a key environmental factor that regulates dormancy and germination of numerous species of plants, including cereals and other Poaceae [34,35]. White light, similarly to blue light, represses the germination of barley, Lolium rigidum, and Brachypodium distachyon, whereas red light stimulates the germination of Brachypodium distachyon but does not affect the dormancy of barley [15,28,[36][37][38].…”