CHARACTERIZING PROTEINS INVOLVED IN B1TR GENE REGULATION

The control of gene expression by cis-regulatory elements is a shared feature across the kingdoms of life. The maize booster1 gene (b1) is a mechanistic model for control of gene expression from a distal cis-regulatory element. Two epialleles of b1 produce distinct tissue-specific pigmentation phenotypes and have been well studied over the past 50 years. The B-Intense (B-I) allele is highly transcribed, while the B’ is lowly transcribed.  When combined in the same the nucleus, B’ paramutates B-I, converting the B-I allele to B’. Expression and paramutation of b1 is influenced by a hepta-tandem repeat sequence (b1TR) located ~100kb upstream of the b1 TSS. Differences in DNA-methylation and chromatin structure has been observed at the b1TR in B’ and B-I plants. It is likely that enhancement and silencing of b1 each rely on distinct trans-factors that may interact with b1TR and modify local chromatin structure. To identify b1TR-interacting proteins, we have developed a discovery-based proteomics approach that utilizes a transgenic copy of b1TR adjacent to a GAL4-upstream activation sequence to capture b1TR chromatin. Chromatin isolated from transgenic B’ and B-I plants will be used as input for chromatin-immunoprecipitation coupled with mass-spectrometry. This approach will allow us to identify b1TR-interacting proteins associated with enhancement, silencing, and paramutation. Preliminary results have identified several proteins that may play a role in b1 regulation by interacting with b1TR.