Regulation of the plant one-carbon metabolic pathway and global gene responses in maize under salt stress

Abstract

One-carbon (C₁) metabolism. C₁ metabolism is central to all organisms, because C₁ units have essential roles in biosyntheses of basic materials for living cells, such as protein, nucleic acids, choline and its derivatives. One unique feature of plant C₁ metabolism is that it channels significant C₁ flux from primary carbon metabolism to methylated metabolites. Part I of this dissertation presents functional analysis of plant methyane-THF reductases (MTHFRs) in Arabidopsis and maize, and regulation of the plant C₁ metabolic pathway and glycine betaine (GlyBet) biosynthesis in maize GlyBet near isogenic lines (NILs). Plant MTHFRs were isolated from Arabidopsis and maize and functionally characterized in yeast. Unlike mammalian MTHFRs, the plant enzymes strongly prefer NADH to NADPH and are not inhibited by S-adenosyl-methionine. An NADH dependent MTHFR reaction could be reversible in the cytosol, supported by radiotracer labeling data using [methyl-¹⁴C]methyltetrahydrofolate. Systematic analyses of profiling transcript abundance of plant C₁ genes using microarrays indicated that the regulation of the plant C₁ metabolic pathway may be "fine-tuned", or at a downstream "outlet" point in maize. Comparison of GlyBet NILs through analyses of metabolites, [¹⁴C]formaldehyde labeling and northern blotting of phosphoethanolamine methyltransferase (P-EAMT) indicated that the regulation glycine betaine biosynthesis is at P-EAMT, whose transcript is probably repressed by its product, phospho-choline. Global gene responses to salt stress. To understand how genes respond to salt stress is very important for the genetic improvement of salt tolerance in plants. Part II of this dissertation presents a systematic analysis of global gene responses to salt stress in maize roots, using microarrays, that contained ∼8,000 ESTs. Microarray expression analysis revealed that 916 different ESTs were up- or down-regulated under salt stress (611 up-regulated, 305 down-regulated), representing 11% of ∼8,000 ESTs printed on slides. These up- or down-regulated ESTs were clustered into various subgroups based on repression or induction of the timing, amplitude and duration of their transcripts. Contig analysis assigned 916 up- or down-regulated ESTs into 472 tentative unique genes (51.6%). These responsive genes are involved in a broad range of cellular processes, biochemical pathways and signal transduction cascades etc. The functions of some regulated genes are discussed.