Minisymposium 16: Gene Regulation

Abs # 28004: Ferredoxin-1 mRNA stability and translation are two separable light regulatory processes conferred by the 5' UTR sequences

Presenter:	Petracek, Marie E, marie_petracek@biochem.okstate.edu
Authors	Petracek, Marie E (A)   Bhat, Sumana  (A)   Krueger, Angela D (A)   Tang, Li  (A)
Affiliations:	(A): Oklahoma State University

Light influences and coordinates gene expression throughout the life of a plant. Genes encoding chloroplast-localized proteins are divided between the nuclear and chloroplast genomes. Following chloroplast maturation, the stoichiometry of these proteins is continually refined through changes in gene expression and protein stability to optimize harvesting light energy according to changing light quality and quantity. Apparent signals from the chloroplast help coordinate gene expression between the two genomes. These signals control nuclear gene expression at the levels of nuclear gene transcription, mRNA stability and cytoplasmic translation. We are using the pea Ferredoxin-1 mRNA (Fed-1) in transgenic tobacco plants as a model for photosynthetic control of mRNA stability and translation. In the light, Fed-1 mRNA is associated with polyribosomes and is stable. In the dark, or in the presence of a photosynthetic electron transport inhibitor, Fed-1 mRNA dissociates from polyribosomes and is destabilized. To identify cis-elements that control Fed-1 mRNA stability and translation, we fused segments of the transcribed portion of Fed-1 to other plant mRNA sequences and found that light-regulated translation of Fed-1 mRNA is controlled by redundant, separable elements within the 5' UTR, coding sequence and the 3' UTR. In contrast, Fed-1 mRNA light-regulated stability is conferred by its 5' UTR. The (CAUU)₄ repeat present in the Fed-1 5' UTR is necessary, but not sufficient, for destabilizing Fed-1 mRNA in dark. Substitution of single nucleotides within this repeat had dramatically different effects on mRNA accumulation. We are currently defining the minimal sequence necessary for dark-induced Fed-1 mRNA destabilization. We have also determined that several portions of the Fed-1 5' UTR are critical for translational regulation. Even in the presence of the redundant 3' UTR and coding sequence elements, mutations upstream of the (CAUU)₄ abolish dark-induced polyribosome dissociation of Fed-1 mRNA, but deletion of this region has no effect on light-regulated translation. We are testing the hypothesis that a transient secondary structure of Fed-1 mRNA disrupts translation in the dark and that mutations upstream of the (CAUU)₄ result in an alternative structure that prohibits formation of the structure necessary for polyribosome dissociation. Also we are identifying proteins that associate with Fed-1 5' UTR in the light and dark using UV-crosslinking assays.