Alleman 9603747 r is one of four loci in maize that regulates the expression of the genes in the anthocyanin biosynthetic pathway through the function of its product, helix-loop-helix transcription factor. r determines the timing and tissue-specificity of anthocyanin deposition during development. By analyzing mutants affecting the regulation of single transcriptonal unit r alleles, the principal investigator has established the basis for investigating tissue-specificity of r. The R(Sc) gene of the R-st complex specifies pigmentation in the aleurone layer of the kernel and the coleoptile and scutellar node of the young seedling. On going studies have identified a 493 bp sequence (B' not) which is located in putative enhancer element in the Sc regulatory region. This segment can be regularly deleted in certain heteroallelic combinations via a proposed gene conversion event with a displaced proximal region to produce enhanced Sc expression Sc|P(enhanced) . Alternatively, the B'not element or the surrounding enhancer region can be disrupted by transposable element Ds mutagenesis. Ds insertions within this region block pigmentation slightly when they are in one orientation or totally when they are in the other orientation, possibly by disrupting the normal domain boundary. The upstream DNA sequence Ds insertional mutants, their derivatives, and a selected group of three variant alleles representing perturbation in strong seed color alleles (Sc) to determine the relationship between the structure and function of one class of r alleles. Using this DNA sequence analysis, it will be determined the mechanism for Sc|P(enhanced) derivative production is by gene conversion. Chromatin structure will also be analyzed for a class of alleles for which pigment is conditionally blocked. An established collection of races of maize differing only by the r allele will be screened using a phenotypic assay and eventually DNA gel blot analysis to locate other related Sc-type alleles. A genetic screen will be region in order to localize other regions that are involved in developmental gene expression. The studies constitute a detailed genetic dissection of the regulatory sequences controlling gene expression in higher plants and are of fundamental importance for understanding how genes are regulated during plant development. By using mutagenesis in which the starting material is a series of variant alleles and single transcriptional unit r alleles determining pigmentation in the same that tissue. The identification of tissue-specific regulatory elements in maize is crucial in order to genetically engineer this agronomically important species.
