Modified physiology of burley tobacco plants genetically engineered to express Yb₁, a functional EGY enzyme

Transgenic overexpression of a NtEGY2 gene restores normal green color of burley tobacco plants, but does not increase nitrogen utilization efficiency beyond that exhibited by wild-type individuals. Nitrogen physiology is important in tobacco because of its role in generation of leaf yield and accumulation of nitrogen-containing alkaloids that can react with nitrosating agents in the formation of carcinogenic tobacco-specific nitrosamines. Cultivars of the burley tobacco market class are homozygous for deleterious mutant alleles at the duplicate Yb₁ and Yb₂ loci which have previously been associated with decreased nitrogen use and utilization efficiency; increased leaf nitrate, total nitrogen, and alkaloid levels; and reduced yields. How mutant alleles at these two loci affect these traits is not well understood. Recent characterization of the Yb₁ and Yb₂ genes (homologs of Arabidopsis EGY1 gene) enabled overexpression of the wild-type Yb₁ allele in yb₁yb₁yb₂yb₂ plants to determine if observed unfavorable effects were due to linkage or pleiotropy, and to determine if overexpression could lead to beneficial modifications in any of these traits in transgenic plants relative to naturally-occurring wild-type genotypes. Yb₁ overexpression was found to confer an agronomic benefit to yb₁yb₁yb₂yb₂ genotypes but no advantage to wild-type genotypes. RNA-Seq was used to carry out a comparative transcriptome analysis of genetically engineered and wild-type nearly isogenic lines (NILs) to gain insight on metabolic pathways affecting carbon and nitrogen metabolism that might be altered as the result of genetic variability at the Yb₁ and Yb₂ loci. Results indicate that complex changes in the transcriptome of tobacco can be manifested by altered expression of Yb₁.