Limits...
Rapid transcriptome responses of maize (Zea mays) to UV-B in irradiated and shielded tissues.

Casati P, Walbot V - Genome Biol. (2004)

Bottom Line: More transcript changes occurred in directly exposed than in shielded organs, and the levels of more transcripts were changed in adult compared to seedling tissues.As the same total UV-B irradiation dose applied at three intensities elicited different transcript profiles, the transcriptome changes exhibit threshold effects rather than a reciprocal dose-effect response.Transcriptome profiling highlights possible signaling pathways and molecules for future research.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biological Sciences, 385 Serra Mall, Stanford University, Stanford, CA 94305-5020, USA. pcasati@stanford.edu

ABSTRACT

Background: Depletion of stratospheric ozone has raised terrestrial levels of ultraviolet-B radiation (UV-B), an environmental change linked to an increased risk of skin cancer and with potentially deleterious consequences for plants. To better understand the processes of UV-B acclimation that result in altered plant morphology and physiology, we investigated gene expression in different organs of maize at several UV-B fluence rates and exposure times.

Results: Microarray hybridization was used to assess UV-B responses in directly exposed maize organs and organs shielded by a plastic that absorbs UV-B. After 8 hours of high UV-B, the abundance of 347 transcripts was altered: 285 were increased significantly in at least one organ and 80 were downregulated. More transcript changes occurred in directly exposed than in shielded organs, and the levels of more transcripts were changed in adult compared to seedling tissues. The time course of transcript abundance changes indicated that the response kinetics to UV-B is very rapid, as some transcript levels were altered within 1 hour of exposure.

Conclusions: Most of the UV-B regulated genes are organ-specific. Because shielded tissues, including roots, immature ears, and leaves, displayed altered transcriptome profiles after exposure of the plant to UV-B, some signal(s) must be transmitted from irradiated to shielded tissues. These results indicate that there are integrated responses to UV-B radiation above normal levels. As the same total UV-B irradiation dose applied at three intensities elicited different transcript profiles, the transcriptome changes exhibit threshold effects rather than a reciprocal dose-effect response. Transcriptome profiling highlights possible signaling pathways and molecules for future research.

Show MeSH

Related in: MedlinePlus

RNA gel-blot analysis to study the kinetics of UV-B induction of gene expression in adult leaves under experimental protocol 2. Lanes contained 10 μg total RNA extracted from adult leaves after 2, 4, 6 or 8 h of UV-B (+) followed by a no UV-B period to complete 8 h, and after 8 h of UV-B followed to a period of 12, 24 or 48 h of no UV-B (+), and from no UV-B (-) treatments. Several identical gels were prepared and blotted. Each blot was hybridized with 32P-labeled clathrin (a) or ribosomal protein L11 (b) probes. (c) Ethidium-bromide-stained gel as a check for equal loading. Bars in gray indicate light treatment without UV-B; bars in white indicate light treatment with UV-B supplementation; and bars in black indicate dark treatment for the time indicated in the figure.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC395766&req=5

Figure 9: RNA gel-blot analysis to study the kinetics of UV-B induction of gene expression in adult leaves under experimental protocol 2. Lanes contained 10 μg total RNA extracted from adult leaves after 2, 4, 6 or 8 h of UV-B (+) followed by a no UV-B period to complete 8 h, and after 8 h of UV-B followed to a period of 12, 24 or 48 h of no UV-B (+), and from no UV-B (-) treatments. Several identical gels were prepared and blotted. Each blot was hybridized with 32P-labeled clathrin (a) or ribosomal protein L11 (b) probes. (c) Ethidium-bromide-stained gel as a check for equal loading. Bars in gray indicate light treatment without UV-B; bars in white indicate light treatment with UV-B supplementation; and bars in black indicate dark treatment for the time indicated in the figure.

Mentions: RNA blot hybridization and real-time RT-PCR were used to analyze the kinetics of UV-B transcript changes in both directly exposed (adult leaf) and shielded (root) tissues. For experiments using adult leaves, two cDNAs that were upregulated within 8 hours in this organ were utilized as probes for northern blots. In the first protocol to determine when transcripts are induced, adult leaves were exposed under UV-B lamps for 2, 4, 6 and 8 hours at 0.36 W/m2; samples were collected immediately after the UV-B treatment from irradiated and control plants. As shown in Figure 8, a 2-hour UV-B exposure suffices to increase transcript levels of clathrin (GenBank AW134461) and ribosomal protein L11 (AI948309), although the increase is lower than the twofold cut-off in the microarray experiments (see Additional data file 1). Clathrin transcripts (Figure 8a) show a progressive increase with longer exposures; in contrast, ribosomal protein L11 transcripts are approximately equivalent at 2 and 8 hours. In the second protocol to explore the persistence of transcript upregulation in the absence of UV-B, leaves were UV-B-irradiated for 2, 4 or 6 hours, followed by a period without UV-B to complete an 8-hour treatment; samples were also collected from plants irradiated for 8 hours followed by a 12-, 24- or 48-hour recovery period. As shown in Figure 9, clathrin and ribosomal protein L11 transcripts are induced with exposures from 2 to 8 hours, as expected from the initial experiment. For clathrin, a short exposure of UV-B is enough to upregulate this gene, but a longer time is required to reach higher levels of expression. Both transcripts persist in recovery periods of 2 hours (after a 6-hour exposure), 6 hours (after a 2-hour exposure), and 12 hours (after an 8-hour exposure). After 12 hours without UV-B, transcripts are lower than at the end of the UV-B treatment, and after 24 hours transcript levels are similar to the non-irradiated control plants. The same RNA samples used in the experiments above were used to compare UV-B-regulated expression of other genes by real-time RT-PCR. Table 2 shows that, as observed for clathrin transcripts, UV-B induction of a cysteine proteinase (GenBank AW129800) shows a progressive increase with longer exposures, while transcripts for a histone deacetylase (AW438666) and for a cytosine 5' DNA methyltransferase (AW215926) are approximately equivalent at 2 and 8 hours.


Rapid transcriptome responses of maize (Zea mays) to UV-B in irradiated and shielded tissues.

Casati P, Walbot V - Genome Biol. (2004)

RNA gel-blot analysis to study the kinetics of UV-B induction of gene expression in adult leaves under experimental protocol 2. Lanes contained 10 μg total RNA extracted from adult leaves after 2, 4, 6 or 8 h of UV-B (+) followed by a no UV-B period to complete 8 h, and after 8 h of UV-B followed to a period of 12, 24 or 48 h of no UV-B (+), and from no UV-B (-) treatments. Several identical gels were prepared and blotted. Each blot was hybridized with 32P-labeled clathrin (a) or ribosomal protein L11 (b) probes. (c) Ethidium-bromide-stained gel as a check for equal loading. Bars in gray indicate light treatment without UV-B; bars in white indicate light treatment with UV-B supplementation; and bars in black indicate dark treatment for the time indicated in the figure.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC395766&req=5

Figure 9: RNA gel-blot analysis to study the kinetics of UV-B induction of gene expression in adult leaves under experimental protocol 2. Lanes contained 10 μg total RNA extracted from adult leaves after 2, 4, 6 or 8 h of UV-B (+) followed by a no UV-B period to complete 8 h, and after 8 h of UV-B followed to a period of 12, 24 or 48 h of no UV-B (+), and from no UV-B (-) treatments. Several identical gels were prepared and blotted. Each blot was hybridized with 32P-labeled clathrin (a) or ribosomal protein L11 (b) probes. (c) Ethidium-bromide-stained gel as a check for equal loading. Bars in gray indicate light treatment without UV-B; bars in white indicate light treatment with UV-B supplementation; and bars in black indicate dark treatment for the time indicated in the figure.
Mentions: RNA blot hybridization and real-time RT-PCR were used to analyze the kinetics of UV-B transcript changes in both directly exposed (adult leaf) and shielded (root) tissues. For experiments using adult leaves, two cDNAs that were upregulated within 8 hours in this organ were utilized as probes for northern blots. In the first protocol to determine when transcripts are induced, adult leaves were exposed under UV-B lamps for 2, 4, 6 and 8 hours at 0.36 W/m2; samples were collected immediately after the UV-B treatment from irradiated and control plants. As shown in Figure 8, a 2-hour UV-B exposure suffices to increase transcript levels of clathrin (GenBank AW134461) and ribosomal protein L11 (AI948309), although the increase is lower than the twofold cut-off in the microarray experiments (see Additional data file 1). Clathrin transcripts (Figure 8a) show a progressive increase with longer exposures; in contrast, ribosomal protein L11 transcripts are approximately equivalent at 2 and 8 hours. In the second protocol to explore the persistence of transcript upregulation in the absence of UV-B, leaves were UV-B-irradiated for 2, 4 or 6 hours, followed by a period without UV-B to complete an 8-hour treatment; samples were also collected from plants irradiated for 8 hours followed by a 12-, 24- or 48-hour recovery period. As shown in Figure 9, clathrin and ribosomal protein L11 transcripts are induced with exposures from 2 to 8 hours, as expected from the initial experiment. For clathrin, a short exposure of UV-B is enough to upregulate this gene, but a longer time is required to reach higher levels of expression. Both transcripts persist in recovery periods of 2 hours (after a 6-hour exposure), 6 hours (after a 2-hour exposure), and 12 hours (after an 8-hour exposure). After 12 hours without UV-B, transcripts are lower than at the end of the UV-B treatment, and after 24 hours transcript levels are similar to the non-irradiated control plants. The same RNA samples used in the experiments above were used to compare UV-B-regulated expression of other genes by real-time RT-PCR. Table 2 shows that, as observed for clathrin transcripts, UV-B induction of a cysteine proteinase (GenBank AW129800) shows a progressive increase with longer exposures, while transcripts for a histone deacetylase (AW438666) and for a cytosine 5' DNA methyltransferase (AW215926) are approximately equivalent at 2 and 8 hours.

Bottom Line: More transcript changes occurred in directly exposed than in shielded organs, and the levels of more transcripts were changed in adult compared to seedling tissues.As the same total UV-B irradiation dose applied at three intensities elicited different transcript profiles, the transcriptome changes exhibit threshold effects rather than a reciprocal dose-effect response.Transcriptome profiling highlights possible signaling pathways and molecules for future research.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biological Sciences, 385 Serra Mall, Stanford University, Stanford, CA 94305-5020, USA. pcasati@stanford.edu

ABSTRACT

Background: Depletion of stratospheric ozone has raised terrestrial levels of ultraviolet-B radiation (UV-B), an environmental change linked to an increased risk of skin cancer and with potentially deleterious consequences for plants. To better understand the processes of UV-B acclimation that result in altered plant morphology and physiology, we investigated gene expression in different organs of maize at several UV-B fluence rates and exposure times.

Results: Microarray hybridization was used to assess UV-B responses in directly exposed maize organs and organs shielded by a plastic that absorbs UV-B. After 8 hours of high UV-B, the abundance of 347 transcripts was altered: 285 were increased significantly in at least one organ and 80 were downregulated. More transcript changes occurred in directly exposed than in shielded organs, and the levels of more transcripts were changed in adult compared to seedling tissues. The time course of transcript abundance changes indicated that the response kinetics to UV-B is very rapid, as some transcript levels were altered within 1 hour of exposure.

Conclusions: Most of the UV-B regulated genes are organ-specific. Because shielded tissues, including roots, immature ears, and leaves, displayed altered transcriptome profiles after exposure of the plant to UV-B, some signal(s) must be transmitted from irradiated to shielded tissues. These results indicate that there are integrated responses to UV-B radiation above normal levels. As the same total UV-B irradiation dose applied at three intensities elicited different transcript profiles, the transcriptome changes exhibit threshold effects rather than a reciprocal dose-effect response. Transcriptome profiling highlights possible signaling pathways and molecules for future research.

Show MeSH
Related in: MedlinePlus