Doctoral Dissertations

Date of Award

5-2002

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Botany

Major Professor

Albrecht Von Arnim

Abstract

Light is one of the most environmental factors that control plant growth and development. Research concerning the effect of light quality and light intensity has focused on the photoreceptors that perceive the light signals, the genetics of photoreceptor-specific signaling pathways, and the developmental responses to light. However little is known about the integration of signals from the various light signaling pathways.

Light signaling pathways and their interactions were investigated in Arabidopsis thaliana by measuring light responsive transcription in a collection of promoter trap lines. Each promoter trap line carries a single, transposon-mediated fusion between a randomly selected endogenous Arabidopsis sequence and the E. coli β-glucuronidase (GUS) reporter gene. Light response profiles, defined as the variation in GUS expression of a gene over a range of environmental conditions, were characterized for a large number of individual promoter traps by histochemical GUS straining and by whole-plant enzyme assays. By applying either constant light or darkness or shifts between light and darkness, I identified 286 light responsive promoter traps. Interestingly, a large fraction of the lines screened displayed tissue-specific light responsiveness. Few examples of this phenomenon have been published based on transgenic promoter:reporter fusions in plants.

The light responsive lines were screened for their response under constant red, constant far-red light, and four types of light pulse regimes in order to define the relative sensitivity of the lines to phytochrome A (phyA) and phytochrome B (phyB) photoreceptors. All tissues surveyed, including the root, were capable of supporting a variety of light response profiles, suggesting that the interaction between light signaling pathways are similarly complex in most cell types. It was very rare to find stronger expression in constant far red than constant red light. This finding suggests that the phyA pathway, active under constant far red, plays a major role in the regulation of genes that are repressed by light and a minor role in genes that are active by light. Because certain light response profiles occurred repeatedly, it was possibly to group the profiles using hierarchical cluster analysis. This revealed groups of genes with antagonistic or synergistic interactions between phyA and phyB.

The chromosomal flanking sequences that drive the light responsive expression profiles were analyzed in detail for fifteen promoter traps. The typical light responsive promoter trap was light repressible, yet contained basal promoter sequence motifs, as well as motifs previously implicated in light inducible gene expression. Many of the light responsive flanking sequences were bona fide cryptic promoters, rather than regulatory regions for native Arabidopsis genes, indicating that cryptic promoters may play an important role in the light-signaling network.

Microarray analyses of individual Arabidopsis cDNAs in response to shifts between light and darkness were performed as a first step to correlate expression profiles identified by promoter trapping with profiles defined at the level of mRNAs. For both light shift experiments, the most commonly activated genes coded for proteins involved in metabolism, and among these, primary metabolism outweighed secondary metabolism. After the shift from constant dark to light, inducible genes preferentially coded for proteins involved in photosynthetic light reactions. In contrast, a shift from light to darkness elicited the expression of genes involved in cell expansion and transport, as expected. More unexpectedly, the shift to darkness also triggered expression of genes involved in translation and stress responses.

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