Germination of photoblastic lettuce seeds is regulated via the control of endogenous physiologically active gibberellin content, rather than of gibberellin responsiveness

Phytochrome regulates lettuce (Lactuca sativa L. cv. Grand Rapids) seed germination via the control of the endogenous level of bioactive gibberellin (GA). In addition to the previously identified LsGA20ox1, LsGA20ox2, LsGA3ox1, LsGA3ox2, LsGA2ox1, and LsGA2ox2, five cDNAs were isolated from lettuce seeds: LsCPS, LsKS, LsKO1, LsKO2, and LsKAO. Using an Escherichia coli expression system and functional assays, it is shown that LsCPS and LsKS encode ent-copalyl diphosphate synthase and ent-kaurene synthase, respectively. Using a Pichia pastoris system, it was found that LsKO1 and LsKO2 encode ent-kaurene oxidases and LsKAO encodes ent-kaurenoic acid oxidase. A comprehensive expression analysis of GA metabolism genes using the quantitative reverse transcription polymerase chain reaction suggested that transcripts of LsGA3ox1 and LsGA3ox2, both of which encode GA 3-oxidase for GA activation, were primarily expressed in the hypocotyl end of lettuce seeds, were expressed at much lower levels than the other genes tested, and were potently up-regulated by phytochrome. Furthermore, LsDELLA1 and LsDELLA2 cDNAs that encode DELLA proteins, which act as negative regulators in the GA signalling pathway, were isolated from lettuce seeds. The transcript levels of these two genes were little affected by light. Lettuce seeds in which de novo GA biosynthesis was suppressed responded almost identically to exogenously applied GA, irrespective of the light conditions, suggesting that GA responsiveness is not significantly affected by light in lettuce seeds. It is proposed that lettuce seed germination is regulated mainly via the control of the endogenous content of bioactive GA, rather than the control of GA responsiveness

A vicious cycle between acid sensing and survival signaling in myeloma cells : acid-induced epigenetic alteration

Myeloma (MM) cells and osteoclasts are mutually interacted to enhance MM growth while creating acidic bone lesions. Here, we explored acid sensing of MM cells and its role in MM cell response to acidic conditions. Acidic conditions activated the PI3K-Akt signaling in MM cells while upregulating the pH sensor transient receptor potential cation channel subfamily V member 1 (TRPV1) in a manner inhibitable by PI3K inhibition. The acid-activated PI3K-Akt signaling facilitated the nuclear localization of the transcription factor Sp1 to trigger the expression of its target genes, including TRPV1 and HDAC1. Consistently, histone deacetylation was enhanced in MM cells in acidic conditions, while repressing a wide variety of genes, including DR4. Indeed, acidic conditions deacetylated histone H3K9 in a DR4 gene promoter and curtailed DR4 expression in MM cells. However, inhibition of HDAC as well as either Sp1 or PI3K was able to restore DR4 expression in MM cells suppressed in acidic conditions. These results collectively demonstrate that acid activates the TRPV1-PI3K-Akt-Sp1 signaling in MM cells while inducing HDAC-mediated gene repression, and suggest that a positive feedback loop between acid sensing and the PI3K-Akt signaling is formed in MM cells, leading to MM cell response to acidic bone lesions

Changes in the transcript levels of gibberellin (GA) metabolism genes in imbibed lettuce seeds after various light treatments

(A) Time-course of lettuce seed germination after light treatment. Zero (0) h indicates seeds imbibed for 3 h in the dark that received no light treatment. FR, FR/R, and FR/R/FR indicate seeds treated with far-red light, far-red followed by red light, and far-red followed by red and then far-red light, respectively. FR/R+ABA indicates seeds treated with far-red followed by red light and 0.1 mM ABA. Triplicate experiments were performed, and means with standard errors are shown. (B) Expression levels of GA metabolism genes after light treatment. The expression levels of these genes were analysed by QRT-PCR. The results were normalized to the expression of 18S rRNA (internal control); the expression levels of all genes examined are given relative to the reference value of the transcript level of at 0 h, set to 1. Three independent experiments were performed, and means with standard errors are shown.<p><b>Copyright information:</b></p><p>Taken from "Germination of photoblastic lettuce seeds is regulated via the control of endogenous physiologically active gibberellin content, rather than of gibberellin responsiveness"</p><p></p><p>Journal of Experimental Botany 2008;59(12):3383-3393.</p><p>Published online 24 Jul 2008</p><p>PMCID:PMC2529229.</p><p></p

Changes in the transcript levels of gibberellin (GA) metabolism genes in the cotyledon end and hypocotyl end of lettuce seeds

(A) Frozen seeds were divided into two parts: cotyledon end, including the cotyledons (Cot), fruit wall (FW), seed coat (SC), and endosperm (ES); and the hypocotyl end, including the hypocotyl (Hyp), root apical meristem (RAM), shoot apical meristem (SAM), and part of the Cot, FW, SC, and ES. (B) Expression levels of GA metabolism genes after light treatment, determined using QRT-PCR. See for light treatments. The results were normalized to the expression of 18S rRNA (internal control), and the highest value was set to 100. Two independent experiments were performed, and means with standard errors are shown.<p><b>Copyright information:</b></p><p>Taken from "Germination of photoblastic lettuce seeds is regulated via the control of endogenous physiologically active gibberellin content, rather than of gibberellin responsiveness"</p><p></p><p>Journal of Experimental Botany 2008;59(12):3383-3393.</p><p>Published online 24 Jul 2008</p><p>PMCID:PMC2529229.</p><p></p

Effects of abscisic acid (ABA) treatment on the expression levels of gibberellin (GA) metabolism genes in imbibed lettuce seeds

(A) The expression levels of the genes were analysed by QRT-PCR. The results were normalized to the expression of 18S rRNA (internal control), and the expression levels of all genes examined are given relative to the reference value of the transcript level of at 0 h, set to 1. Three independent experiments were performed, and means with standard errors are shown. (B) Expression analysis using seeds that had been cut in half. The results were normalized to the expression of 18S rRNA (internal control), and the highest value was set to 100. Two independent experiments were performed, and means with standard errors are shown.<p><b>Copyright information:</b></p><p>Taken from "Germination of photoblastic lettuce seeds is regulated via the control of endogenous physiologically active gibberellin content, rather than of gibberellin responsiveness"</p><p></p><p>Journal of Experimental Botany 2008;59(12):3383-3393.</p><p>Published online 24 Jul 2008</p><p>PMCID:PMC2529229.</p><p></p

Expression of and during germination, and gibberellin (GA) responsiveness in lettuce seeds

(A) Expression levels of and after light treatment. The expression analysis was carried out by QRT-PCR. See for light treatments. The results were normalized to the expression of 18S rRNA (internal control), and the expression levels of all genes examined are given relative to the reference value of the transcript level of at 0 h, set to 1. Three independent experiments were performed, and means with standard errors are shown. (B) Expression levels of and in the cotyledon end and the hypocotyl end of lettuce seeds after light treatment. The results were normalized to the expression of 18S rRNA (internal control), and the highest value was set to 100. Two independent experiments were performed, and means with standard errors are shown. (C) Germination frequency of lettuce seeds in the presence of a GA biosynthesis inhibitor and various concentrations of GA. Five sets of 20 decoated lettuce seeds were incubated in the dark at 25 °C in medium containing 50 μM uniconazol-P and various concentrations of GA. After red light treatment, seeds were incubated at 25 °C in the dark for 24 h and the germination frequency was recorded. Means with standard errors are shown.<p><b>Copyright information:</b></p><p>Taken from "Germination of photoblastic lettuce seeds is regulated via the control of endogenous physiologically active gibberellin content, rather than of gibberellin responsiveness"</p><p></p><p>Journal of Experimental Botany 2008;59(12):3383-3393.</p><p>Published online 24 Jul 2008</p><p>PMCID:PMC2529229.</p><p></p