Imbibition, but Not Release from Stratification, Sets the Circadian Clock in Arabidopsis Seedlings

Abstract

Circadian rhythms in the abundance of the CAT2 catalase mRNA were not seen in etiolated seedlings but developed upon illumination. These circadian oscillations were preceded by a rapid and transient induction of CAT2 mRNA abundance that varied strikingly according to the timing (circadian phase) of the onset of illumination. This variation oscillated with a circadian periodicity of ‫ف‬ 28 hr, indicating that the circadian oscillator is running in etiolated seedlings and regulates (gates) the induction of CAT2 by light. Moreover, because we assayed populations of seedlings, we infer that the individual clocks among populations of etiolated seedlings were synchronized before the onset of illumination. What developmental or environmental signals synchronized the clocks among seedlings? Varying the phase of the onset of illumination relative to release from stratification failed to affect the acute induction of CAT2 , indicating that the temperature step from 4 to 22 ؇ C associated with release from stratification did not reset the circadian clock. However, the acute induction of CAT2 mRNA varied with time after imbibition, demonstrating that imbibition provides a signal capable of resetting the circadian clock and of synchronizing the clocks among populations of seedlings. INTRODUCTIONThe biological circadian clock generates circadian rhythms with periods of ‫ف‬ 24 hr. Plants are richly rhythmic, and the clock provides endogenous timing information that is used to coordinate numerous aspects of plant development and physiology (Sweeney, 1987;Kreps and Kay, 1997;Lumsden and Millar, 1998). The circadian clock regulates the expression of many plant genes that serve as molecular markers of the status of the clock. These circadian rhythms are not unique to plants but are widespread, if not ubiquitous, among eukaryotes and prokaryotes (Edmunds, 1988; Dunlap, 1996; Golden et al., 1997).The catalase ( CAT ) gene family of Arabidopsis provides a particularly useful set of molecular markers for the circadian clock because the individual members of this small family of three genes each respond differently to temporal information supplied by the circadian clock (McClung, 1997). Although CAT1 mRNA abundance does not exhibit circadian oscillation (Frugoli et al., 1996), there are circadian oscillations in the mRNA abundance of both CAT2 and CAT3. It is intriguing that the peaks in mRNA abundance of CAT2 and CAT3 are gated to distinct phases (times of day) by the circadian clock, with CAT2 mRNA being most abundant at dawn and CAT3 mRNA being most abundant at dusk (Zhong et al., 1994;Zhong and McClung, 1996). The physiological significance of the circadian regulation of CAT mRNA abundance is not yet apparent. Catalase (H 2 O 2 :H 2 O 2 oxidoreductase; EC 1.11.1.6) belongs to a group of enzymes involved in regulating the cellular levels of active oxygen species. Catalase is present in all aerobic organisms and protects cells from the damaging effects of H 2 O 2 by converting H 2 O 2 to O 2 and H 2 O. However, it remains to be dete...

Integrated Temporal Regulation of the Photorespiratory Pathway. Circadian Regulation of Two Arabidopsis Genes Encoding Serine Hydroxymethyltransferase

“…When etiolated seedlings were transferred into continuous light, SHM1 mRNA accumulated to a peak about 24 h after the onset of illumination and then declined slightly, which is reminiscent of the pattern observed with transcripts such as LHCB and CAT2, which are regulated by the circadian clock (Tavladoraki et al, 1989;Zhong et al, 1994Zhong et al, , 1998Millar and Kay, 1996). To directly assess whether a circadian clock controls SHM1 and SHM4 mRNA abundance, we measured mRNA abundance in whole seedlings germinated and grown for 7 d in 12-h light:12-h dark cycles and then released into continuous light.…”

“…A circadian clock also regulates the mRNA abundance of the photorespiratory catalase gene CAT2 (Zhong et al, 1994), a peroxisomal component of the photorespiratory pathway. Moreover, a circadian clock gates the response of CAT2 to environmental stimuli such as light (Zhong et al, 1998). To determine whether a circadian clock coordinates the temporal expression of mitochondrial components of the photorespiratory pathway with these chloroplastic and peroxisomal components, we set out to characterize the SHM genes encoding Ser hydroxymethyltransferase, an essential mitochondrial matrix component of the photorespiratory pathway.…”

“…In contrast, both phytochromes A and B contribute to the acute induction of Arabidopsis LHCB (CAB2) mRNA by light (Karlin-Neumann et al, 1988;Anderson et al, 1997). The transfer of etiolated seedlings into continuous white light induces circadian oscillations in LHCB (Tavladoraki et al, 1989;Millar and Kay, 1996) and CATALASE (Zhong et al, 1994(Zhong et al, , 1998 mRNAs. It is likely that the accumulation of SHM1 mRNA observed after 24 h of illumination with white, red, or blue light represents sampling during the peak of the circadian oscillation in SHM1 mRNA, because the phase of the oscillation of SHM1 mRNA is similar to the phases of the oscillations in LHCB and CATALASE 2 mRNAs, both of which peak about 24 h after the onset of illumination.…”

“…Light-regulated genes such as LHCB and CAT2 exhibit an acute response to light within about 4 h of the onset of illumination (Zhong et al, 1994(Zhong et al, , 1998Millar and Kay, 1996). In contrast, SHM1 did not exhibit an acute induction, suggesting that the accumulation of SHM1 mRNA in the light is an indirect response to illumination, and may represent a consequence of the stimulation of photosynthesis and photorespiration in the light.…”

GIGANTEA Regulates Phytochrome A-Mediated Photomorphogenesis Independently of Its Role in the Circadian Clock

“…To synchronize the rhythms, the seeds were sown at 7 PM local time because imbibition sets the clock to an evening phase (Zhong et al, 1998). The seeds were incubated 61 h in darkness at 5°C and given 11 h R to induce germination and returned to darkness (22°C), i.e.…”