Plants have acquired and maintained an expanded suite of DNA-dependent RNA polymerases (RNAPs) compared to other eukaryotes. Although their exact roles remain unclear, plant-specific RNAPs (Pol IV and Pol V) are involved in epigenetic silencing of transposable elements (TEs). Zea mays (maize) Pol IV is required for proper plant development as well as the establishment and maintenance of paramutations, which are trans-homolog interactions that facilitate heritable gene silencing. Maize has duplications of Pol IV catalytic subunits, which define multiple Pol IV subtypes, and accessory proteins associated with these subtypes define distinct Pol IV complexes. Understanding the roles of Pol IV will require identifying the composition and function of these Pol IV subtypes and complexes. In exploring interactions between the genes encoding a Pol IV catalytic subunit and a putative accessory protein, I identified two new alleles of the Pol IV subunit and a family of potential Pol IV accessory proteins conserved in multicellular plants. Together, these Pol IV complexes are proposed to function through two, potentially overlapping, general mechanisms to control of gene function: direct competition with Pol II and/or generation of 24-nucleotide RNAs that guide de novo cytosine methylation. I analyzed nascent transcriptome data of seedlings lacking Pol IV and their wild-type siblings, which identified a global effect of Pol IV on gene boundary transcription. Pol IV-affected loci serve as molecular and phenotypic models for dissecting the involvement of various Pol IV subtype and complex components. Such analysis implicated a Pol IV-affected allele as being controlled by Pol IV competition with Pol II. Through expanded sequencing of a paramutation participating allele (Pl1-Rhoades), I identified a new Pol IV target of five tandem repeats that may serve as an enhancer element. I used the Pl1-Rhoades allele in particular to compare and contrast the effects of Pol IV loss with that of Pol IV accessory proteins. Together, the work presented here explores the involvement of an uncharacterized protein family of putative Pol IV accessory proteins in Pol IV complexes, expands the roles of Pol IV complexes in controlling maize gene function, and identifies new Pol IV-affected loci for future study.
|Advisor:||Hollick, Jay B., Rine, Jasper D.|
|Commitee:||Eisen, Michael B., Freeling, Michael R.|
|School:||University of California, Berkeley|
|Department:||Molecular & Cell Biology|
|School Location:||United States -- California|
|Source:||DAI-B 79/09(E), Dissertation Abstracts International|
|Subjects:||Botany, Genetics, Bioinformatics|
|Keywords:||Nascent transcriptome, Paramutation, Pol IV, RNA polymerases, Rddm, Zea mays|
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