Fluorescent Differential Display Identifies Circadian Clock-Regulated Genes in Arabidopsis thaliana

Abstract

Circadian rhythms in gene expression were first observed in plants more than 13 years ago, but the underlying mechanism controlling rhythmic gene expression is still not understood. The isolation of novel circadian clock-controlled genes (ccgs) is likely to provide new tools for studying circadian rhythms. Fluorescent differential display (FDD) was used to screen Arabidopsis thaliana mRNAs for cycling transcripts. Seventy PCR primer pairs were screened, and 17 different cycling bands were observed out of an estimated 10,500 bands screened. The identities of 10 bands were determined, and the rhythmic gene expression was confirmed using northern blot analysis. The 10 cycling bands represent 7 different genes, 6 of which are present in the databases and 1 that does not match anything in current databases. The rhythmic expression of the 7 genes is composed of four distinct phases of clock regulation. The results demonstrate that FDD can be used to isolate ccgs. The genes identified in this screen range from known A. thaliana ccgs, as well as genes shown to be clock controlled in other plant species, to a novel gene that may encode a pioneer protein. Further study of these ccgs is likely to increase our understanding of circadian-regulated gene expression.

Cloning and Functional Characterization of a β-Pinene Synthase from Artemisia annua That Shows a Circadian Pattern of Expression

“…Functional genomic studies of Arabidopsis show many genes to be controlled by the clock, and their functions oscillate rhythmically (Harmer et al, 2000;Kreps et al, 2000;Schaffer et al, 2001).…”

Chronopharmacognosy

“…It is possible that the timing of expression of such genes (for instance, the predawn rise in Lhcb ), indicates a role for the clock in the coordination of metabolism to maximize photosynthetic yields. The use of fluorescent differential display[19] and high density Deoxyribonucleic acid (DNA) arrays to monitor global expression profiles should give us an indication of the range of the genes showing circadian control.Cytosolic concentrations of free calcium have been shown to oscillate with a circadian rhythm in Arabidopsis. Considering the importance of calcium as both a secondary messenger and a cofactor for many enzymes, this might be a means by which the clock regulates a variety of cellular process.…”

“…It is possible that the timing of expression of such genes (for instance, the predawn rise in Lhcb ), indicates a role for the clock in the coordination of metabolism to maximize photosynthetic yields. The use of fluorescent differential display[19] and high density Deoxyribonucleic acid (DNA) arrays to monitor global expression profiles should give us an indication of the range of the genes showing circadian control.…”

Circadian rhythms in Arabidopsis: time for nuclear proteins

“…It has often been noted coincidentally that many light-regulated genes also continued to disappear and reappear at circadian intervals in constant light (Kloppstech 1985). Subsequently, deliberate isolation of oscillating transcripts was done by systematic subtractive hybridisation of timed cDNA libraries or dierential display (Heintzen et al 1994a, b;Kreps et al 2000).…”